Identification of staphylococci by pyrolysis gas-liquid chromatography.
The identification of bacteria by pyrolysis gas-liquid chromatography (PGLC) was evaluated with the genus Staphylococcus. A matrix containing the characteristics of seven species of Staphylococcus was constructed from the results obtained with 42 named strains identified by biochemical methods. An identification program based upon this database was evaluated in tests with 100 isolates of catalase-positive oxidase-negative gram-positive cocci identified by biochemical and PGLC systems. Agreement between the two systems was 90%. Pure cultures of unknown isolates were identified within 30 min by PGLC using a microcomputer data acquisition and analysis program. A taxonomic analysis of the data is presented, and methods for quality control of PGLC, prolongation of column life, and data analysis are described and discussed.


How do root fungi of Alnus nepalensis and Schima wallichii recover during succession of abandoned land?
Alnus nepalensis and Schima wallichii are native tree species accompanying succession in abandoned agricultural land in the middle mountainous region of central Nepal. To understand how root fungi recover during spontaneous succession, we analyzed the diversity and composition of arbuscular mycorrhizal (AM), ectomycorrhizal (ECM), and total fungi in tree fine roots from three land use types, short-term abandoned land (SA), long-term abandoned land (LA), and regenerated forest (RF) as a reference. Additionally, ECM morphotypes were examined. The results showed different speeds of succession in the studied fungal groups. While the change in the AM fungal community appears to be rapid and LA resembles the composition of RF, the total fungi in the abandoned land types are similar to each other but differed significantly from RF. Interestingly, the relative abundance of Archaeosporaceae followed a trend differing between the tree species (SA < LA in A. nepalensis, but SA > LA in S. wallichii). Unlike AM and total fungi, there was no significant difference in the ECM community of A. nepalensis between land use types, probably due to their low species diversity (9 ECM morphotypes, 31 ECM operational taxonomic units). However, Cortinarius sp. was significantly more abundant in RF than in the other land use types, whereas Alnicola, Tomentella, and Russula preferred young stages. Our results suggest that for both studied tree species the AM fungal succession could reach the stage of regenerated forest relatively fast. In the case of total fungi, because of hyperdiversity and composed of species specialized to a variety of environments and substrates, the transition was expected to be delayed in abandoned land where the vegetation was still developing and the ecosystem was not as complex as that found in mature forests.


[Photogeneration of a 2-vector transmembrane proton gradient in Halobacterium halobium R1 cells].
Analysing 4 phases of light-dependent change of pH in cell suspension of H. halobium R1 it has been found that an increase of pH I when light is switched on and a decrease of pH II during further illumination are proportional to the light effect and are energy-dependent. Neutralization of these changes (phases III and IV) proceeds spontaneously in the darkness. These data show that the transmembrane gradient of protons is generated in two directions--delta pH and +delta pH, simultaneous presence of which points to the local character of one of them. Generation of -delta pH associated with light energy utilization by the cells is considered as a result of neutralization of the negative charge and +delta pH1 (latent), appearing due to a change in bacteriorodopsin conformation under illumination. It may be also suggested that the yield of protons from the cells of pHII and generation of +delta pH result from the discoupling of photoprocesses and energy utilization by the cells, containing bacteriorodopsin.


The coarse-grained plaque: a divergent Aβ plaque-type in early-onset Alzheimer's disease.
Alzheimer's disease (AD) is characterized by amyloid-beta (Aβ) deposits, which come in myriad morphologies with varying clinical relevance. Previously, we observed an atypical Aβ deposit, referred to as the coarse-grained plaque. In this study, we evaluate the plaque's association with clinical disease and perform in-depth immunohistochemical and morphological characterization. The coarse-grained plaque, a relatively large (Ø ≈ 80 µm) deposit, characterized as having multiple cores and Aβ-devoid pores, was prominent in the neocortex. The plaque was semi-quantitatively scored in the middle frontal gyrus of Aβ-positive cases (n = 74), including non-demented cases (n = 15), early-onset (EO)AD (n = 38), and late-onset (LO)AD cases (n = 21). The coarse-grained plaque was only observed in cases with clinical dementia and more frequently present in EOAD compared to LOAD. This plaque was associated with a homozygous APOE ε4 status and cerebral amyloid angiopathy (CAA). In-depth characterization was done by studying the coarse-grained plaque's neuritic component (pTau, APP, PrPC), Aβ isoform composition (Aβ40, Aβ42, AβN3pE, pSer8Aβ), its neuroinflammatory component (C4b, CD68, MHC-II, GFAP), and its vascular attribution (laminin, collagen IV, norrin). The plaque was compared to the classic cored plaque, cotton wool plaque, and CAA. Similar to CAA but different from classic cored plaques, the coarse-grained plaque was predominantly composed of Aβ40. Furthermore, the coarse-grained plaque was distinctly associated with both intense neuroinflammation and vascular (capillary) pathology. Confocal laser scanning microscopy (CLSM) and 3D analysis revealed for most coarse-grained plaques a particular Aβ40 shell structure and a direct relation with vessels. Based on its morphological and biochemical characteristics, we conclude that the coarse-grained plaque is a divergent Aβ plaque-type associated with EOAD. Differences in Aβ processing and aggregation, neuroinflammatory response, and vascular clearance may presumably underlie the difference between coarse-grained plaques and other Aβ deposits. Disentangling specific Aβ deposits between AD subgroups may be important in the search for disease-mechanistic-based therapies.


Dietary and lifestyle characteristics of colorectal cancer in Jordan: a case-control study.
We evaluated dietary pattern and lifestyle characteristics of patients with colorectal cancer (CRC) in Jordan. The case-control study included 220 recently diagnosed CRC cases and 220 age and gender matched healthy subjects as a control group. The participating CRC cases had lower dietary intake of fibre, folate, vitamin B12, β-carotene, vitamin C and selenium as compared to controls (P<0.05). The frequency of consumption of fruits and vegetables was also lower among CRC cases, while the frequency of consumption of red meat and saturated fat was higher and positively associated with CRC risk. Furthermore, family history for CRC played a positive role and the majority of CRC cases and controls had a low physical activity level. A sedentary lifestyle and a diet low in fruits and vegetables, and high in animal red meat and saturated fat, appeared associated with CRC among the studied Jordanian subjects. This is consistent with the reported CRC studies in developed nations indicating global causal effects for this tumour type.


Use of a Gabapentin Protocol for the Management of Alcohol Withdrawal: A Preliminary Experience Expanding From the Consultation-Liaison Psychiatry Service.
Benzodiazepines are the conventional mainstay to manage alcohol withdrawal; however, patients are subsequently at increased risk for poor sleep, cravings, and return to drinking. Research on alternative pharmacologic agents to facilitate safe alcohol withdrawal is scant. Gabapentin is one medication shown in small studies to reduce the need for benzodiazepines in the setting of alcohol withdrawal. The continuation of gabapentin after alcohol withdrawal appears to be safe during early sobriety and may aid in reducing alcohol-related cravings or returning to alcohol consumption. Use of a gabapentin-based, benzodiazepine-sparing protool began in early 2015 by the Mayo Clinic, Rochester, Consultation-Liaison Psychiatry Service. A retrospective chart review was conducted to detect any safety concerns with use of a gabapentin protocol for alcohol withdrawal syndrome. Secondary outcomes were derived by comparing a matched cohort of patients who received benzodiazepines for alcohol withdrawal syndrome. Seventy-seven patients had their alcohol withdrawal managed via a gabapentin protocol during the study period. No patients required transfer to a higher level of care or had a documented withdrawal seizure. Length of stay between the gabapentin protocol group and benzodiazepine group were similar. This preliminary data has supported the frequent use of this protocol in the general internal medicine practice and formalization of an institutional order set of this protocol for mild to moderate alcohol withdrawal syndrome. Prospective studies are required to validate findings.


Brazilian Green Propolis Extract Synergizes with Protoporphyrin IX-mediated Photodynamic Therapy via Enhancement of Intracellular Accumulation of Protoporphyrin IX and Attenuation of NF-κB and COX-2.
Brazilian green propolis (BGP) is noted for its impressive antitumor effects and has been used as a folk medicine in various cultures for many years. It has been demonstrated that BGP could enhance the cytotoxic effect of cytostatic drugs on tumor cells. Photodynamic therapy (PDT) is a therapeutic approach used against malignant cells. To assess the synergistic effect of BGP extract on protoporphyrin IX (PpIX)-mediated photocytotoxicity, MTT assays were performed using A431 and HeLa cells. TUNEL assay and Annexin V-FITC/PI staining were performed to confirm the induction of apoptosis. Western blotting analysis was performed to examine the pro-apoptotic proteins, anti-apoptotic proteins and inflammation related proteins in A431 cells. Intracellular accumulation of PpIX was examined by flow cytometry. The synergistic effect of BGP extract in PpIX-PDT was also evaluated with a xenograft model. Our findings reveal that BGP extract increased PpIX-mediated photocytotoxicity in A431 and HeLa cells. PpIX-PDT with BGP extract treatment resulted in a decrease in Bcl-xL and an increase in NOXA, Bax and caspase-3 cleavage. The protein expression levels of p-IKKα/β, NF-κB and COX-2 were upregulated by PpIX-PDT but significantly attenuated when in combination with BGP extract. BGP extract was also found to significantly enhance the intracellular accumulation of PpIX in A431 cells. BGP extract increased PpIX-mediated photocytotoxicity in a xenograft model as well. Our findings provide evidence for a synergistic effect of BGP extract in PpIX-PDT both in vitro and in vivo.


Noninvasive assessment of the developing Xenopus cardiovascular system using optical coherence tomography.
Studies investigating normal and abnormal cardiac development are frequently limited by an inability to assess cardiovascular function within the intact organism. In this work, optical coherence tomography (OCT), a new method of micron-scale, noninvasive imaging based on the measurement of backscattered infrared light, was introduced for the high resolution assessment of structure and function in the developing Xenopus laevis cardiovascular system. Microstructural details, such as ventricular size and wall positions, were delineated with OCT at 16-microm resolution and correlated with histology. Three-dimensional representation of the cardiovascular system also was achieved by repeated cross-sectional imaging at intervals of 25 microm. In addition to structural information, OCT provides high speed in vivo axial ranging and imaging, allowing quantitative dynamic activity, such as ventricular ejection fraction, to be assessed. The sensitivity of OCT for dynamic assessment was demonstrated with an inotropic agent that altered cardiac function and dimensions. Optical coherence tomography is an attractive new technology for assessing cardiovascular development because of its high resolution, its ability to image through nontransparent structures, and its inexpensive portable design. In vivo and in vitro imaging are performed at a resolution approaching that of histopathology without the need for animal killing.


Forensic aspects of mass disasters: strategic considerations for DNA-based human identification.
Many mass disasters result in loss of lives. Law enforcement and/or public safety and health officials often have the responsibility for identifying the human remains found at the scene, so they can be returned to their families. The recovered human remains range from being relatively intact to highly degraded. DNA-based identity testing is a powerful tool for victim identification in that the data are not restricted to any particular one to one body landmark comparison and DNA profile comparisons can be used to associate separated remains or body parts. Even though DNA typing is straightforward, a disaster is a chaotic environment that can complicate effective identification of the remains. With some planning, or at least identification of the salient features to consider, stress can be reduced for those involved in the identification process. General guidelines are provided for developing an action plan for identification of human remains from a mass disaster by DNA analysis. These include: (1) sample collection, preservation, shipping and storage; (2) tracking and chain of custody issues; (3) laboratory facilities; (4) quality assurance and quality control practices; (5) parsing out work; (6) extraction and typing; (7) interpretation of results; (8) automation; (9) software for tracking and managing data; (10) the use of an advisory panel; (11) education and communication; and (12) privacy issues. In addition, key technologies that may facilitate the identification process are discussed, such as resin based DNA extraction, real-time PCR for quantitation of DNA, use of mini-STRs, SNP detection procedures, and software. Many of the features necessary for DNA typing of human remains from a mass disaster are the same as those for missing persons' cases. Therefore, developing a missing persons DNA identification program would also provide the basis for a mass disaster human remains DNA identification program.


Phenotypic and genotypic characterization of Staphylococci causing breast peri-implant infections in oncologic patients.
Staphylococcus epidermidis and S. aureus have been identified as the most common bacteria responsible for sub-clinical and overt breast implant infections and their ability to form biofilm on the implant as been reported as the essential factor in the development of this type of infections. Biofilm formation is a complex process with the participation of several distinct molecules, whose relative importance in different clinical settings has not yet been fully elucidated. To our knowledge this is the first study aimed at characterizing isolates causing breast peri-implant infections. Thirteen S. aureus and seven S. epidermidis causing breast peri-implant infections were studied. Using the broth microdilution method and the E-test, the majority of the strains were susceptible to all antibiotics tested. Methicillin resistance was detected in two S. epidermidis. All strains had different RAPD profiles and were able to produce biofilms in microtitre plate assays but, while all S. aureus carried and were able to express icaA and icaD genes, this was only true for one S. epidermidis. Biofilm development was glucose- and NaCl-induced (5 S. aureus and 1 S. epidermidis) or glucose-induced (the remaining strains). Proteinase K and sodium metaperiodate treatment had different effects on biofilms dispersion revealing that the strains studied were able to produce chemically different types of extracellular matrix mediating biofilm formation. All S. aureus strains harboured and expressed the atlA, clfA, FnA, eno and cna genes and the majority also carried and expressed the sasG (10/13), ebpS (10/13) genes. All S. epidermidis strains harboured and expressed the atlE, aae, embp genes, and the majority (six strains) also carried and expressed the fbe, aap genes. Genes for S. aureus capsular types 5 and 8 were almost equally distributed. The only leukotoxin genes detected were lukE/lukD (6/13). S. aureus and S. epidermidis breast peri-implant infections are caused by heterogeneous strains with different biofilm development mechanisms. Since the collagen adhesin (cna) gene is not ubiquitously distributed among S. aureus, this protein could have an important role in the cause of breast peri-implant infections.


Copper chaperone Atox1 interacts with the metal-binding domain of Wilson's disease protein in cisplatin detoxification.
Human copper transporters ATP7B (Wilson's disease protein) and ATP7A (Menkes' disease protein) have been implicated in tumour resistance to cisplatin, a widely used anticancer drug. Cisplatin binds to the copper-binding sites in the N-terminal domain of ATP7B, and this binding may be an essential step of cisplatin detoxification involving copper ATPases. In the present study, we demonstrate that cisplatin and a related platinum drug carboplatin produce the same adduct following reaction with MBD2 [metal-binding domain (repeat) 2], where platinum is bound to the side chains of the cysteine residues in the CxxC copper-binding motif. This suggests the same mechanism for detoxification of both drugs by ATP7B. Platinum can also be transferred to MBD2 from copper chaperone Atox1, which was shown previously to bind cisplatin. Binding of the free cisplatin and reaction with the cisplatin-loaded Atox1 produce the same protein-bound platinum intermediate. Transfer of platinum along the copper-transport pathways in the cell may serve as a mechanism of drug delivery to its target in the cell nucleus, and explain tumour-cell resistance to cisplatin associated with the overexpression of copper transporters ATP7B and ATP7A.


Efficient virus-induced gene silencing in roots using a modified tobacco rattle virus vector.
Due to their capability of eliciting a form of posttranscriptional gene silencing (termed virus-induced gene silencing or VIGS), plant viruses are increasingly used as reverse-genetics tools for functional characterization of plant genes. RNA viruses have been shown to trigger silencing in a variety of host plants, including members of Solanacae and Arabidopsis (Arabidopsis thaliana). Several factors affect the silencing response, including host range and viral tropism within the plant. The work presented here demonstrates that a modified tobacco rattle virus (TRV) vector retaining the helper protein 2b, required for transmission by a specific vector nematode, not only invades and replicates extensively in whole plants, including meristems, but also triggers a pervasive systemic VIGS response in the roots of Nicotiana benthamiana, Arabidopsis, and tomato (Lycopersicon esculentum). This sustained VIGS response was exemplified by the silencing of genes involved in root development (IRT1, TTG1 [transparent testa glabra], RHL1 [root hairless1], and beta-tubulin), lateral root-meristem function (RML1 [root meristemless1]), and nematode resistance (Mi). Roots of silenced plants exhibit reduced levels of target mRNA and phenocopy previously described mutant alleles. The TRV-2b vector displays increased infectivity and meristem invasion, both key requirements for efficient VIGS-based functional characterization of genes in root tissues. Our data suggest that the TRV helper protein 2b may have an essential role in the host regulatory mechanisms that control TRV invasion.


Blueberry Yields Increase With Bee Visitation Rates, but Bee Visitation Rates are not Consistently Predicted by Colony Stocking Densities.
Pollinator-dependent crops rely on the activity of managed and wild pollinators. While farm management and surrounding landscape can influence wild pollinator contributions, managed pollinator contributions may be primarily driven by their stocking densities, though this is not well studied across crops. We selected 20 southern highbush blueberry farms along two independent gradients of honey bee Apis mellifera L. (Hymenoptera: Apidae) stocking density (~1-11 hives/acre) and bumble bee Bombus impatiens Cresson (Hymenoptera: Apidae) stocking density (0 - 3 colonies/acre) ensuring that stocking densities were not correlated with farm or landscape attributes. Across farms, we observed managed and wild bee visitation rates, and measured yield estimates. Farms with greater bumble bee stocking densities had higher bumble bee visitation rates and yield estimates, but farms with higher honey bee stocking densities only received higher honey bee visitation rates at the end of bloom and did not have higher yield estimates. The main wild pollinator, the southeastern blueberry bee Habropoda laboriosa (Fabricius) (Hymenoptera: Apidae), showed higher visitation rates on organic farms and in late bloom. In general, higher visitation rates by honey bees, bumble bees, and H. laboriosa were correlated with higher yields. Our results suggest that yields are limited by bee visitation rates, and that within the stocking density ranges studied, increasing managed bumble bees, but not honey bees, increases their visitation rates. While H. laboriosa had the greatest effect on yield estimates, its activity appears to be limited by both a phenological mismatch with crop bloom and farm management.


Changes in total nitrogen, soluble protein, and peroxidases in the expanding leaf zone of eastern cottonwood.
Nitrogen content and soluble protein and anodal peroxidase banding in acrylamide gel changed with leaf and internode development in the expanding leaf zone of eastern cottonwood (Populus deltoides Bartr.). Nitrogen per unit leaf area was high near the apex and decreased to a constant value at the sixth node below it. Soluble protein banding was qualitatively similar for leaves and internodes in this zone, but anodal peroxidases differed between leaves and internodes. The major leaf peroxidase band was absent from the second leaf below the apex but present in the fourth and sixth leaves; its appearance and intensification seemed to parallel the development of photosynthetic activity. The major internode peroxidase band was present in the apex, second, fourth, and sixth internodes, and intensified during internodal development. It is suggested that these two "isoenzymes" may have different functions in vivo.


Maturational differences in the isolated isovolumic rabbit heart.
To determine maturational differences in myocardial pressure work, oxygen consumption, and stability, we evaluated an isolated retrogradely perfused isovolumic rabbit heart model in newborns (0-6 days of age, mean 3.3 +/- 1.9 days), immatures (4-6 wk) and adults (5-7 mo). To reduce ischemic time and cardiac trauma, the hearts were isolated and instrumented within the chest cavity. By design, there were no significant age-related differences in coronary flow per gram, heart rate, or resting pressure. Left ventricular developed pressure was similar at all ages, but myocardial oxygen consumption was much lower in newborns and immatures than in adults. Coronary resistance was lower in the younger hearts. There were no significant age-related differences in contractility, left ventricular stiffness, or myocardial pH. All hearts were stable for 120 min, but thereafter immatures and adults deteriorated more rapidly than newborns. The study demonstrates that this model is technically suitable for the study of age-related differences in cardiac physiology. Furthermore, newborn and immature hearts are able to more efficiently use oxygen to develop left ventricular pressures, possibly due to a lower wall tension.


Looking Outside: Summer Camp for Children Who Have Undergone a Solid-Organ Transplant.
A high-quality critical care team is an essential component of any successful organ transplant program. From pretransplant care to the crucial postoperative period, its importance cannot be discounted. However, because of the focused nature of work in an intensive care unit (ICU), all too often members of the ICU team are not able to see and appreciate the ultimate fruits of their labor. These are factors that can contribute to the high rates of burnout and turnover among ICU teams. This article presents the concept of a summer camp for children who have received a solid-organ transplant. We discuss a vehicle by which ICU staff as well as other members of a patient's care team can gain a better appreciation of the full nature of both medical and, perhaps more importantly in this situation, nonmedical goals and outcomes in pediatric transplantation. We review our institutional experience running a summer camp for these children, discuss some of the important points in making such a camp successful, and discuss potential benefits to the campers as well as those taking care of them.


Charting the relationship trajectories of aggressive, withdrawn, and aggressive/withdrawn children during early grade school.
The premises examined in this longitudinal investigation were that specific behavioral characteristics place children at risk for relationship maladjustment in school environments, and that multiple behavioral risks predispose children to the most severe and prolonged difficulties. Aggressive, withdrawn, and aggressive/withdrawn children were compared to normative and matched control groups on teacher and peer relationship attributes, loneliness, and social satisfaction from kindergarten (M age = 5 years, 7 months; n = 250) through grade 2 (M age = 8.1; n = 242). Children's withdrawn behavior was neither highly stable nor predictive of relational difficulties, as their trajectories resembled the norm except for initially less close and more dependent relationships with teachers. Aggressive behavior was fairly stable, and associated with early-emerging, sustained difficulties including low peer acceptance and conflictual teacher-child relationships. Aggressive/withdrawn children evidenced the most difficulty: compared to children in the normative group, they were consistently more lonely, dissatisfied, friendless, disliked, victimized, and likely to have maladaptive teacher-child relationships. Findings are discussed with respect to recent developments in two prominent literatures: children at-risk and early relationship development.


Constant Electricity Generation in Nanostructured Silicon by Evaporation-Driven Water Flow.
Recently, hydrovoltaic technology emerged as a novel renewable energy harvesting method, which dramatically extends the capability to harvest water energy. However, the urgent issue restricting its device performance is poor carrier transport properties of the solid surface if large charged interface is considered simultaneously. Herein, a hydrovoltaic device based on silicon nanowire arrays (SiNWs), which provide large charged surface/volume ratio and excellent carrier transport properties, yields sustained electricity by a carrier concentration gradient induced by evaporation-induced water flow inside nanochannels. The device can yield direct current with a short-circuit current density of over 55 μA cm-2 , which is three orders larger than a previously reported analogous device (approximately 40 nA cm-2 ). Moreover, it exhibits a constant output power density of over 6 μW cm-2 and an open-circuit voltage of up to 400 mV. Our finding may pave a way for developing energy-harvesting devices from ubiquitous evaporation-driven internal water flow in nature with semiconductor material of silicon.


Diffusion weighted imaging in cystic fibrosis disease: beyond morphological imaging.
To explore the feasibility of diffusion-weighted imaging (DWI) to assess inflammatory lung changes in patients with Cystic Fibrosis (CF) METHODS: CF patients referred for their annual check-up had spirometry, chest-CT and MRI on the same day. MRI was performed in a 1.5 T scanner with BLADE and EPI-DWI sequences (b = 0-600 s/mm2). End-inspiratory and end-expiratory scans were acquired in multi-row scanners. DWI was scored with an established semi-quantitative scoring system. DWI score was correlated to CT sub-scores for bronchiectasis (CF-CTBE), mucus (CF-CTmucus), total score (CF-CTtotal-score), FEV1, and BMI. T-test was used to assess differences between patients with and without DWI-hotspots. Thirty-three CF patients were enrolled (mean 21 years, range 6-51, 19 female). 4 % (SD 2.6, range 1.5-12.9) of total CF-CT alterations presented DWI-hotspots. DWI-hotspots coincided with mucus plugging (60 %), consolidation (30 %) and bronchiectasis (10 %). DWItotal-score correlated (all p < 0.0001) positively to CF-CTBE (r = 0.757), CF-CTmucus (r = 0.759) and CF-CTtotal-score (r = 0.79); and negatively to FEV1 (r = 0.688). FEV1 was significantly higher (p < 0.0001) in patients without DWI-hotspots. DWI-hotspots strongly correlated with radiological and clinical parameters of lung disease severity. Future validation studies are needed to establish the exact nature of DWI-hotspots in CF patients. • DWI hotspots only partly overlapped structural abnormalities on morphological imaging • DWI strongly correlated with radiological and clinical indicators of CF-disease severity • Patients with more DWI hotspots had lower lung function values • Mucus score best predicted the presence of DWI-hotspots with restricted diffusion.


Sources of variation in cell-type RNA-Seq profiles.
Cell-type specific gene expression profiles are needed for many computational methods operating on bulk RNA-Seq samples, such as deconvolution of cell-type fractions and digital cytometry. However, the gene expression profile of a cell type can vary substantially due to both technical factors and biological differences in cell state and surroundings, reducing the efficacy of such methods. Here, we investigated which factors contribute most to this variation. We evaluated different normalization methods, quantified the variance explained by different factors, evaluated the effect on deconvolution of cell type fractions, and examined the differences between UMI-based single-cell RNA-Seq and bulk RNA-Seq. We investigated a collection of publicly available bulk and single-cell RNA-Seq datasets containing B and T cells, and found that the technical variation across laboratories is substantial, even for genes specifically selected for deconvolution, and this variation has a confounding effect on deconvolution. Tissue of origin is also a substantial factor, highlighting the challenge of using cell type profiles derived from blood with mixtures from other tissues. We also show that much of the differences between UMI-based single-cell and bulk RNA-Seq methods can be explained by the number of read duplicates per mRNA molecule in the single-cell sample. Our work shows the importance of either matching or correcting for technical factors when creating cell-type specific gene expression profiles that are to be used together with bulk samples.


Cost shifting: new myths, old confusion, and enduring reality.
The papers by Paul Ginsburg and Jason Lee and colleagues make it clear how far we have come on the issue of cost shifting and how far we still have to go. We have created some new myths, persisted in some old confusion, and largely ignored an enduring reality. That enduring reality is that lack of competition in the health care sector permits cost shifting to occur. Instead of focusing on this causal condition, public discussion has been centered on the symptoms. A number of steps can be taken to deal with causes; this paper discusses several of these steps.


Genotoxicity of novel trans-platinum(II) complex with diethyl (pyridin-4-ylmethyl)phosphate in human non-small cell lung cancer cells A549.
The dynamic development of metal-containing anticancer drugs has started since the discovery of cis-diamminedichloroplatinum(II). For many years it was believed that trans platinum(II) compounds were non-active as antitumour agents because trans-diamminedichloroplatinum is biologically inactive although it binds to DNA and also forms monoadducts and cross-links. In the present work the ability of a novel platinum(II) compound trans-[PtCl(2)(4-pmOpe)(2)] to induce DNA damage in human non-small cell lung cancer cells A549 was examined using the alkaline comet assay. The obtained results revealed that the novel trans platinum(II) complex induced DNA strand breaks, which were effectively repaired during 2h of post-incubation, and cross-links which remained unrepaired under these test conditions. Apart from that, the modified comet assay with incubation with proteinase K was used to verify the ability of trans-[PtCl(2)(4-pmOpe)(2)] and cis-DDP to form DNA-protein cross-links. It has been proved that only trans-[PtCl(2)(4-pmOpe)(2)] complex exhibits the ability to induce DNA-protein cross-links. The results suggest a different mechanism of action of this compound in comparison to cis-DDP. It seems that trans geometry and the presence of two diethyl (pyridin-4-ylmethyl)phosphates as non-leaving ligands can determine dissimilar properties of the adducts formed on DNA and the different mechanism of action of trans-[PtCl(2)(4-pmOpe)(2)] and in consequence the efficacy in killing cancer cells.


Winged helix transcription factor Foxb1 is essential for access of mammillothalamic axons to the thalamus.
Our aim was to study the mechanisms of brain histogenesis. As a model, we have used the role of winged helix transcription factor gene Foxb1 in the emergence of a very specific morphological trait of the diencephalon, the mammillary axonal complex. Foxb1 is expressed in a large hypothalamic neuronal group (the mammillary body), which gives origin to a major axonal bundle with branches to thalamus, tectum and tegmentum. We have generated mice carrying a targeted mutation of Foxb1 plus the tau-lacZ reporter. In these mutants, a subpopulation of dorsal thalamic ventricular cells "thalamic palisade" show abnormal persistence of Foxb1 transcriptional activity; the thalamic branch of the mammillary axonal complex is not able to grow past these cells and enter the thalamus. The other two branches of the mammillary axonal complex (to tectum and tegmentum) are unaffected by the mutation. Most of the neurons that originate the mammillothalamic axons suffer apoptosis after navigational failure. Analysis of chimeric brains with wild-type and Foxb1 mutant cells suggests that correct expression of Foxb1 in the thalamic palisade is sufficient to rescue the normal phenotype. Our results indicate that Foxb1 is essential for diencephalic histogenesis and that it exerts its effects by controlling access to the target by one particular axonal branch.


Pharmacogenetic Interventions Improve the Clinical Outcome of Treatment-Resistant Autistic Spectrum Disorder Sufferers.
Autistic spectrum disorders (ASD) are severe neurodevelopmental alterations characterised by deficits in social communication and repetitive and restricted behaviours. About a third of patients receive pharmacological treatment for comorbid symptoms. However, 30-50% do not respond adequately and/or present severe and long-lasting side effects. Genetic variants in CYP1A2, CYP2C19, CYP2D6 and SLC6A4 were investigated in N = 42 ASD sufferers resistant to pharmacological treatment. Clinical recommendations based on their pharmacogenetic profiles were provided within 24-48 h of receiving a biological sample. A total of 39 participants (93%) improved after the pharmacogenetic intervention according to their CGI scores (difference in basal-final scores: 2.26, SD 1.55) and 37 participants (88%) according to their CGAS scores (average improvement of 20.29, SD 11.85). Twenty-three of them (55%) achieved symptom stability (CGI ≤ 3 and CGAS improvement ≥ 20 points), requiring less frequent visits to their clinicians and hospital stays. Furthermore, the clinical improvement was higher than that observed in a control group (N = 62) with no pharmacogenetic interventions, in which 66% responded to treatment (difference in CGI scores: -0.87, SD 9.4, p = 1 × 10-5; difference in CGAS scores: 6.59, SD 7.76, p = 5 × 10-8). The implementation of pharmacogenetic interventions has the potential to significantly improve the clinical outcomes in severe comorbid ASD populations with drug treatment resistance and poor prognosis.


GPR30: A new potential therapeutic target in human testicular germ cell tumors.
The G protein-coupled estrogen receptor (GPR30) is suggested to exert a role in non-nuclear estrogen signalling and is over-expressed in a variety of hormone dependent tumors. It is well known that estrogens and xenoestrogens are involved in testicular germ cell tumorigenesis. Different studies show that down regulation of estrogen receptor β (ERβ) associates with GPR30 over-expression both in human testicular carcinoma in situ (CIS) and seminomas and that the mitogenic role exerted by 17β-oestradiol induces the activation of extracellular signal-regulated kinase 1/2 (ERK1/2) through GPR30. In conclusion, the exposure to oestrogens or oestrogen-mimics, in some as of yet undefined manner, diminishes the ERβ-mediated growth restraint in CIS and in human testicular seminoma, indicating that GPR30 could be considered a potential therapeutic target to design specific inhibitors.


Microwave, high-resolution infrared, and quantum chemical investigations of CHBrF2: ground and v4 = 1 states.
A combined microwave, infrared, and computational investigation of CHBrF(2) is reported. For the vibrational ground state, measurements in the millimeter- and sub-millimeter-wave regions for CH(79)BrF(2) and CH(81)BrF(2) provided rotational and centrifugal-distortion constants up to the sextic terms as well as the hyperfine parameters (quadrupole-coupling and spin-rotation interaction constants) of the bromine nucleus. The determination of the latter was made possible by recording of spectra at sub-Doppler resolution, achieved by means of the Lamb-dip technique, and supporting the spectra analysis by high-level quantum chemical calculations at the coupled-cluster level. In this context, the importance of relativistic effects, which are of the order of 6.5% and included in the present work using second-order direct perturbation theory, needs to be emphasized for accurate predictions of the bromine quadrupole-coupling constants. The infrared measurements focused on the ν(4) fundamental band of CH(79)BrF(2). Fourier transform investigations using a synchrotron radiation source provided the necessary resolution for the observation and analysis of the rotational structure. The spectroscopic parameters of the v(4) = 1 state were found to be close to those of the vibrational ground state, indicating that the ν(4) band is essentially unaffected by perturbations.


The evolving roles of nuclear cardiology.
The use of cardiac imaging modalities has grown steadily, and cardiac nuclear studies constitute a large part of this number. Nuclear Cardiology is often mistakenly considered a synonym of myocardial perfusion imaging (MPI), but has broader applications, including metabolic imaging, innervation imaging, among other technologies. MPI has been a powerful diagnostic and prognostic tool in the assessment of patients for known or suspected CAD for decades, and is now increasingly used for the evaluation of the anti-ischemic effects of various therapies, according to changes in left ventricular perfusion defect size defined by sequential MPI. Neuronal dysfunction identified with iodine-123-metaiodobenzylguanidine may give information on prognosis in different disease conditions, such as after myocardial infarction, in diabetes and dilated cardiomyopathy. Molecular imaging may identify the predominant cellular population in the atherosclerotic plaque and help predict the likelihood of clinical events. Therefore, although its usefulness is well established, Nuclear Cardiology remains a moving science, whose roles keep in pace with evolving clinical needs and expectations.


Central nervous system insulin receptors in normal and diabetic rats.
The presence of specific insulin receptors in homogenates of various parts of the rat brain was demonstrated. Hypothalamic preparations exhibited the greatest insulin binding. Scatchard plots of the binding data were typically curvilinear. Insulin binding to the brain homogenates was influenced by pH, incubation time, temperature, and ionic milieu. The specificity of insulin binding to the brain homogenates was demonstrated by partial displacement of labeled insulin by insulin analogs according to their biological activities and by inhibition of insulin binding by serum containing an antibody to insulin receptors. Insulin binding to brain homogenates from streptozotocin-induced diabetic rats was similar to that from control animals and failed to indicate up-regulation of these receptors.


Too Little Exercise and Too Much Sitting: Inactivity Physiology and the Need for New Recommendations on Sedentary Behavior.
Moderate- to vigorous-intensity physical activity has an established preventive role in cardiovascular disease, type 2 diabetes, obesity, and some cancers. However, recent epidemiologic evidence suggests that sitting time has deleterious cardiovascular and metabolic effects that are independent of whether adults meet physical activity guidelines. Evidence from "inactivity physiology" laboratory studies has identified unique mechanisms that are distinct from the biologic bases of exercising. Opportunities for sedentary behaviors are ubiquitous and are likely to increase with further innovations in technologies. We present a compelling selection of emerging evidence on the deleterious effects of sedentary behavior, as it is underpinned by the unique physiology of inactivity. It is time to consider excessive sitting a serious health hazard, with the potential for ultimately giving consideration to the inclusion of too much sitting (or too few breaks from sitting) in physical activity and health guidelines.


Graphene and novel graphitic ZnO and ZnS nanofilms: the energy landscape, non-stoichiometry and water dissociation.
The energy landscapes of ultra-thin nanofilms of ZnO and ZnS are examined in detail using periodic hybrid density functional calculations. We predict new staggered graphitic forms, which are stable only for the thinnest films and are of particular interest as the electronic structure shows a spontaneous symmetry breaking across the film and consequently a marked decrease in band gap with thickness. The relative energies of the various forms, their structural and electronic properties and their variation with film thickness are discussed. Possible kinetic pathways for transitions from the graphitic forms are examined by explicit evaluation of transition state energies. For polar surfaces, such as (0001) würtzite and (111) zinc blende, many different mechanisms operate to remove or reduce the surface dipole depending on the number of layers in the nanofilm. The polar ZnS nanofilms, but not the polar ZnO analogues or any non-polar film, are predicted to spontaneously become non-stoichiometric by loss of zinc atoms from the surface. The behaviour of adsorbed water on the ultra-thin films is also examined. There is no dissociation on any ZnS film. For ZnO, dissociation into OH- and H+ takes place not only on (101̄0) würtzite, but also on (110) zinc blende. This result that does not appear to have been reported previously and deserves future experimental study. While we concentrate on ZnO and ZnS, similar energy landscapes are expected for any oxide or sulphide which adopts the würtzite or zinc blende structure in the bulk.


Addenbrooke's Cognitive Examination-Revised for mild cognitive impairment in Parkinson's disease.
Cognitive impairment is common in Parkinson's disease (PD), even in the early stages, and appropriate screening tools are needed. We investigated the utility of the Addenbrooke's Cognitive Examination-Revised for detecting mild cognitive impairment (MCI) in PD in an incident population-representative cohort (n = 132) and investigated the relationship between performance on this instrument and behavior and quality of life (n = 219). Twenty-two percent met criteria for MCI. Receiver operating curve analysis revealed an area under the curve of 0.81. A cutoff <89 gave a sensitivity of 69% and specificity of 84%. Scores on this instrument were highly correlated with the Parkinson's Disease Cognitive Rating Scale, and there were significant correlations with the Cambridge Behavioral Inventory-Revised and Parkinson's Disease Questionnaire 39. This instrument is a useful screening tool for PD-MCI, and poor performance is significantly related to impaired behavior and quality of life.


Scrutinizing the FTO locus: compelling evidence for a complex, long-range regulatory context.
Single nucleotide polymorphisms (SNPs) within a genetic region including the first two introns of the gene encoding FTO have consistently been shown to be the strongest genetic factors influencing body mass index (BMI). However, this same also contains several regulatory DNA elements that affect the expression of IRX3 and IRX5, which respectively, are located approximately 500 kb and 1.2 Mbp downstream from the BMI-associated FTO locus. Through these affected regulatory elements, genetic variation at the FTO locus influences adipocyte development leading to decreased thermogenesis and increased lipid storage. These findings provide a genomic model for the functional implications of genetic variations at this locus, and also demonstrate the importance of accounting for chromatin-chromatin interactions when constructing hypotheses for the mechanisms of trait and disease-associated common genetic variants. Several consortia have generated genome-wide datasets describing different aspects of chromatin biology which can be utilized to predict functionality and propose biologically relevant descriptions of specific DNA regions. Here, we review some of the publically available data resources on genome function and organization that can be used to gain an overview of genetic regions of interest and to generate testable hypotheses for future studies. We use the BMI- and obesity-associated FTO locus as a subject as it poses an illustrative example on the value of these resources. We find that public databases strongly support long-range interactions between regulatory elements in the FTO locus with the IRXB cluster genes IRX3 and IRX5. Chromatin configuration capture data also support interactions across a large region stretching across from the RPGRIP1L gene, FTO and the IRXB gene cluster.


Analysis of Putative Epigenetic Regulatory Elements in the FXN Genomic Locus.
Friedreich´s ataxia (FRDA) is an autosomal recessive disease caused by an abnormally expanded Guanine-Adenine-Adenine (GAA) repeat sequence within the first intron of the frataxin gene (FXN). The molecular mechanisms associated with FRDA are still poorly understood and most studies on FXN gene regulation have been focused on the region around the minimal promoter and the region in which triplet expansion occurs. Nevertheless, since there could be more epigenetic changes involved in the reduced levels of FXN transcripts, the aim of this study was to obtain a more detailed view of the possible regulatory elements by analyzing data from ENCODE and Roadmap consortia databases. This bioinformatic analysis indicated new putative regulatory regions within the FXN genomic locus, including exons, introns, and upstream and downstream regions. Moreover, the region next to the end of intron 4 is of special interest, since the enhancer signals in FRDA-affected tissues are weak or absent in this region, whilst they are strong in the rest of the analyzed tissues. Therefore, these results suggest that there could be a direct relationship between the absence of enhancer sequences in this specific region and their predisposition to be affected in this pathology.


Retroperitoneoscopic Heminephrectomy in Duplex Kidney in Infants and Children: Results of a Multicentric Survey.
Retroperitoneoscopic partial nephrectomy (RPN) in children is considered a complex technique with limited diffusion among pediatric surgeons and urologists. We aimed to report the outcome of this technique in infants and children with duplex kidney in a 5-year retrospective multicentric international survey. Data on 50 children who underwent RPN (41 upper-pole nephrectomies and 9 lower-pole nephrectomies) were retrospectively collected in this six-institution survey. Median age at surgery was 3.3 years. There were 35 girls and 15 boys. The left side was affected in 28 patients, versus the right side in 22 patients. We assessed intraoperative and postoperative morbidity. Follow-up (median, 2.5 years; range, 12 months-5 years) was based on clinical controls and echo color Doppler renal ultrasound scans. Median duration of surgery was 255 minutes. Surgery was always performed with the patient in a lateral position. Special hemostatic devices were used for dissection and parenchymal section in all centers. Three patients from two centers (6%) required conversion to open surgery. We recorded seven complications (six peritoneal perforations, one opening of the remaining calyxes) in the 50 cases. Re-operation rate was 0%. Average length of hospital stay was 4.1 days. Our survey shows that RPN remains a challenging procedure with a long learning curve, performed only in pediatric centers with huge experience in this field. In our survey operative time was longer than 4 hours. The complication rate remains high (7/50, or 14%), with complications classified as Grade II according to the Clavien-Dindo classification. They did not require further surgery, but they were associated with a prolonged hospital stay.


Metabolism of glycyrrhetic acid by rat liver microsomes: glycyrrhetinate dehydrogenase.
Glycyrrhetic acid, derived from a main component of liquorice, was converted to 3-ketoglycyrrhetic acid reversibly by rat liver homogenates in the presence of NADPH or NADP+. Glycyrrhetic acid-oxidizing and 3-ketoglycyrrhetic acid-reducing activities were localized in microsomes among the subcellular fractions of rat liver. Glycyrrhetic acid-oxidizing activity and 3-ketoglycyrrhetic acid-reducing activities showed pH optima at 6.3 and 8.5, respectively, and required NADP+ or NAD+ and NADPH or NADH, respectively, indicating that these activities were due to glycyrrhetinate dehydrogenase. The dehydrogenase was not solubilized from the membranes by the treatment with 1 M NaCl or sonication, indicating that the enzyme is a membrane component. The dehydrogenase was solubilized with detergents such as Emalgen 913, Triton X-100 and sodium cholate, and then separated from 3 beta-hydroxysteroid dehydrogenase (5 beta-androstan-3 beta-ol-17-one-oxidizing activity) by butyl-Toyopearl 650 M column chromatography. Partially purified enzyme catalyzed the reversible reaction between glycyrrhetic acid and 3-ketoglycyrrhetic acid, but was inactive toward 3-epiglycyrrhetic acid and other steroids having the 3 beta-hydroxyl group. The enzyme required NADP+ and NADPH for the highest activities of oxidation and reduction, respectively, and NAD+ and NADH for considerable activities, similar to the results with microsomes. From these results the enzyme is defined as glycyrrhetinate dehydrogenase, being quite different from 3 beta-hydroxysteroid dehydrogenase of Ruminococcus sp. from human intestine, which is active for both glycyrrhetic acid and steroids having the 3 beta-hydroxyl group.


Removal of toluene vapors from the polluted air with modified natural zeolite and titanium dioxide nanoparticles.
Toluene is a colorless and flammab1le liquid with the same solubilizing capacity as benzene that is in many cases used as an alternative to benzene, because of the uncertainty of being carcinogens. Workers can be exposed to toluene by breathing the chemical. To avoid inhalation and dermal effects caused by exposure to toluene, solutions such as adsorption, thermal oxidation, membrane separation and photocatalytic processes are applied. In this study, removal of toluene vapors with modified natural zeolite and titanium dioxide nanoparticles was discussed. The natural zeolite was modified using chemical and thermal methods. The samples characterized by Brunauer-Emmet-Teller, Barrett-Joyner-Halenda, X-ray diffraction and scanning electron microscopy tests. After stabilization of TiO2 nanoparticles, the removal efficiency for the toluene vapors at a concentration of 50, 150 and 300 ppm were evaluated using a dynamic system. The results showed that the zeolite has a very porous surface and after modifying the context its specific surface area increased 2.54 times. The results of the adsorption capacity calculation and photocatalytic process showed that modified zeolite samples-TiO2 bed has greater efficiency in the adsorption capacity and better photocatalytic activity than a Ze-TiO2 bed. Ze-TiO2 bed was able to remove 26% of toluene vapors at the concentration of 50 ppm and inlet flow rate of 1 L/m, which was 1.26 times more than a bed Ze-TiO2. According to the results of this study, while modifying the natural zeolite increased desirable properties such as specific surface area and Si/Al ratio, but in comparison with similar studies with synthetic bed such as zeolite Y and ZSM-5, could not achieve desired results in a photocatalytic activity for its application in industry. However, because of its abundance in the world and Iran and therefore low cost of preparation and also due to its unique characteristics, it is recommended that more studies to be done about modifying and its application in photocatalytic processes.


Prediction of 8-year risk of cardiovascular diseases in Korean adult population.
Although many prediction models for cardiovascular diseases (CVDs) have been developed and validated for Western populations, the development of CVD prediction models for Asians has been slow. Our cohort study retrospectively analyzed the incidence of CVD that occurred between January 1, 2009, and December 31, 2016, in all Koreans who underwent national health screening. This dataset included 21,581,796 adults between the ages of 40 and 79 years (10,412,947 men, 11,168,849 women) without CVD at baseline. The primary outcome, CVD, was defined as the development of any of the following: acute coronary syndrome, cerebral infarction, and cerebral hemorrhage, as defined with health insurance claims data. The prediction model was constructed by Cox proportional hazard regression and validated with tenfold cross-validation. The performance of the models was evaluated through Harrell's C-index and Brier score. The discrimination of the models was assessed by the area under the receiver operating characteristic curve (AUROC). Our model showed an AUROC of 0.762 in men and 0.811 in women. The Brier score of our model was 0.018 in men and 0.010 in women, which was better than the pooled cohort equation (PCE). Our novel model performed better than the FRS and PCE for Koreans.


NATIONAL HIV TESTING CAMPAIGNS TO SUPPORT UNAIDS 90-90-90 AGENDA: A LESSON FROM KENYA.
HIV diagnosis is the gateway to antiretroviral therapy. However, 20-50% of HIV-infected individuals are unaware of their HIV status, derailing epidemic control. To increase awareness of HIV status and enrollment into HIV care & treatment (C&T) services through a national HIV testing services (HTS) rapid results initiative (RRI) campaign in Kenya. This cross-sectional analysis presents yield of undiagnosed people living with HIV (PLHIV) and their enrollment into HIV C&T resulting from HTS RRI implemented in July-August 2013 as an example of utilizing RRIs to catalyze achievement of UNAIDS targets. During the campaign 1,462,378 persons received HTS, of whom 220,902 (15%) were children (aged <15 years), 55,088 (7%) couples and 116,126 (8%) key populations. A total of 37,630 (2.6%) HIV+ individuals were identified. Among children who received HTS, 3,244 (1.5%) tested HIV positive, compared to 34,386 (2.8%) among adults. Of the eight regions in Kenya: Nyanza, Rift-valley and Nairobi contributed 73.3% of all HIV+ individuals identified. HTS at health facility settings yielded the highest proportion (69%) of HIV+ and key populations had the highest prevalence (4.8%). Of those infected, 29,851 (79.3%) were enrolled into HIV C&T. Sex, age and setting of HTS were significantly associated with enrollment into HIV C&T (p<0.0001). National HTS campaigns have the potential of increasing knowledge of HIV status. Targeted provision of HTS at health facility settings, to key populations and high burden geographical regions would narrow the gap of undiagnosed PLHIV towards achieving UNIADS 90-90-90 targets for HIV epidemic control.


Dynamic Task Optimization in Remote Diabetes Monitoring Systems.
Diabetes is the seventh leading cause of death in the United States, but careful symptom monitoring can prevent adverse events. A real-time patient monitoring and feedback system is one of the solutions to help patients with diabetes and their healthcare professionals monitor health-related measurements and provide dynamic feedback. However, data-driven methods to dynamically prioritize and generate tasks are not well investigated in the domain of remote health monitoring. This paper presents a wireless health project (WANDA) that leverages sensor technology and wireless communication to monitor the health status of patients with diabetes. The WANDA dynamic task management function applies data analytics in real-time to discretize continuous features, applying data clustering and association rule mining techniques to manage a sliding window size dynamically and to prioritize required user tasks. The developed algorithm minimizes the number of daily action items required by patients with diabetes using association rules that satisfy a minimum support, confidence and conditional probability thresholds. Each of these tasks maximizes information gain, thereby improving the overall level of patient adherence and satisfaction. Experimental results from applying EM-based clustering and Apriori algorithms show that the developed algorithm can predict further events with higher confidence levels and reduce the number of user tasks by up to 76.19 %.


Multi-Parameter Laser Imaging Reveals Complex Microscale Biofilm Matrix in a Thick (4,000 μm) Aerobic Methanol Oxidizing Community.
Although methanol has frequently been used as an inexpensive supplementary carbon source to support treatment processes, knowledge of the resultant microbial biofilms, their 3D architecture, microenvironments, exopolymer chemistry and populations remains limited. We supplied methanol as a supplementary carbon source to biofilms developing in rotating annular reactors. Analysis of circulation waters (1.0 l d-1) indicated that dissolved organic carbon was reduced by 25%, NO3-nitrogen by 95%, and total phosphorus by 70%. Analyses of populations using culture based techniques and fluorescence in situ hybridization indicated enrichment of nitrifiers, denitrifiers, and methylotrophic bacteria relative to reference biofilms not receiving methanol. The biofilms that developed were up to 4,000 μm thick. Staining with fluor conjugated lectins in combination with nucleic acid stains, revealed the presence of discrete bacterial cells inside complex globular polymeric structures. These structures were in turn surrounded by an interstitial polymer containing a variety of bacterial cell types. The globular structures bound FITC-conjugated lectins, from Canavalia ensiformis and Ulex europeaus. The FITC-lectin of Phaseolus vulgaris bound the surface of the globular structures and more generally within the matrix. Chemical analyses of the polymer paralleled the results of lectin analyses indicating that the dominant neutral sugars were glucose, galactose, mannose, rhamnose, with fucose and ribose as minor constituents. Amino sugars were not detected. Dual channel imaging with pH sensitive probes indicated that pH gradients from pH 4 to 7 occurred across the globular microcolonies. Critically for the maintenance of aerobic conditions throughout the thick biofilm it was extensively penetrated by a fine fissure network revealed by the location of fluorescent latex microbeads as detected by confocal laser scanning microscopy. Microelectrode studies confirmed the absence of any detectable Eh gradients within the biofilm. However, mobility of various size-fractionated fluorescent probes indicated that the basal region was only penetrated by the lowest molecular weight probes with a hydrated radius of 2.2 nm or less. These observations indicate the selection of a unique, thick (>4,000 μm) microbial community in which a self-organized architecture promotes the maintenance of optimal conditions and metabolism throughout the biofilm community.


Comparison of Isolated Lumbar Extension Strength in Competitive and Noncompetitive Powerlifters, and Recreationally Trained Men.
Androulakis-Korakakis, P, Gentil, P, Fisher, JP, and Steele, J. Comparison of isolated lumbar extension strength in competitive and noncompetitive powerlifters, and recreationally trained men. J Strength Cond Res 35(3): 652-658, 2021-Low-back strength has been shown to significantly impact performance in a plethora of sports. Aside from its effect on sport performance, low-back strength is strongly associated with low-back pain. A sport that heavily involves the lower-back musculature is powerlifting. This study looked to compare isolated lumbar extension (ILEX) strength in competitive and noncompetitive powerlifters, and recreationally trained men. Thirteen competitive powerlifters (CPL group; 31.9 ± 7.6 years; 173.4 ± 5.5 cm; 91.75 ± 18.7 kg), 10 noncompetitive powerlifters (NCPL group; 24 ± 3.5 years; 179 ± 4.8 cm; 92.39 ± 15.73 kg), and 36 recreationally trained men (RECT group; 24.9 ± 6.5 years; 178.5 ± 5.2 cm; 81.6 ± 10.0 kg) were tested for ILEX. Isolated lumbar extension strength was measured at every 12° throughout subject's full range of motion (ROM) and expressed as the following: "strength index (SI)" calculated as the area under a torque curve from multiple angle testing, average torque produced across each joint angle (AVG), and maximum torque produced at a single angle (MAX). Deadlift and squat strength were measured using 1 repetition maximum for the competitive and noncompetitive powerlifters. The following powerlifting characteristics were recorded for the competitive and noncompetitive powerlifters: primary deadlift stance, primary squat bar position, use of belt, use of performance-enhancing drugs, and use of exercises to target the lower-back musculature. Significant between-group effects were found for subject characteristics (age, stature, body mass, and ROM). However, analysis of covariance with subject characteristics as covariates found no significant between-group effects for SI (p = 0.824), AVG (p = 0.757), or MAX (p = 0.572). In conclusion, this study suggests that powerlifting training likely has little impact on conditioning of the lumbar extensors.


scMD: cell type deconvolution using single-cell DNA methylation references.
The proliferation of single-cell RNA sequencing data has led to the widespread use of cellular deconvolution, aiding the extraction of cell type-specific information from extensive bulk data. However, those advances have been mostly limited to transcriptomic data. With recent development in single-cell DNA methylation (scDNAm), new avenues have been opened for deconvolving bulk DNAm data, particularly for solid tissues like the brain that lack cell-type references. Due to technical limitations, current scDNAm sequences represent a small proportion of the whole genome for each single cell, and those detected regions differ across cells. This makes scDNAm data ultra-high dimensional and ultra-sparse. To deal with these challenges, we introduce scMD (single cell Methylation Deconvolution), a cellular deconvolution framework to reliably estimate cell type fractions from tissue-level DNAm data. To analyze large-scale complex scDNAm data, scMD employs a statistical approach to aggregate scDNAm data at the cell cluster level, identify cell-type marker DNAm sites, and create a precise cell-type signature matrix that surpasses state-of-the-art sorted-cell or RNA-derived references. Through thorough benchmarking in several datasets, we demonstrate scMD's superior performance in estimating cellular fractions from bulk DNAm data. With scMD-estimated cellular fractions, we identify cell type fractions and cell type-specific differentially methylated cytosines associated with Alzheimer's disease.


"I did it without hesitation. Am I the bad guy?" Online conversations in response to controversial in-game violence.
Video game content has evolved over the last six decades, from a basic focus on challenge and competition to include more serious and introspective narratives capable of encouraging critical contemplation within gamers. The "No Russian" mission from Call of Duty: Modern Warfare 2 casts players as terrorists responsible for the murder of innocent bystanders, sparking debate around how players engage and react to wanton violence in modern video games. Through thematic analysis of 649 Reddit posts discussing the mission, 10 themes emerged representing complexity in player experiences. Those themes were grouped into categories representing (descending order), (1) rote gameplay experiences, (2) dark humor, (3) comparing the mission to other games and real-world events, and (4) self-reflective eudaimonic reactions to the mission. Although less common, the presence of eudaimonic media effects (in at least 15% of posts) holds promise for the use of video games as reflective spaces for violence prevention.


Hepatitis B and liver cancer beliefs among Korean immigrants in Western Washington.
Hepatocellular cancer occurs more frequently among Koreans, Vietnamese, and Chinese than other racial/ethnic groups in the U.S. This excess risk can be attributed to high rates of chronic hepatitis B viral (HBV) infection and low rates of HBV vaccination among Asian immigrants. However, there is little available information regarding the hepatitis B knowledge, beliefs, and practices among Koreans, the fifth-largest Asian population in the U.S. This brief report summarizes results from 30 qualitative interviews and two focus groups investigating hepatitis and liver cancer prevention, behavior, and beliefs among first-generation Korean immigrant adults ages 18-64 years residing in the Seattle-Tacoma metropolitan area of Washington State. The report concludes with suggestions for future investigations to address the high rates of chronic HBV infection and hepatocellular cancer in this vulnerable population.


Biochemical characterization of purified, human recombinant Lys304-->Glu medium-chain acyl-CoA dehydrogenase containing the common disease-causing mutation and comparison with the normal enzyme.
Recombinant, normal human medium-chain acyl-CoA dehydrogenase (MCADH) and the common, human disease-causing K304E mutant ([Glu304]MCADH) protein were expressed in Escherichia coli using an optimized system, and the enzymes were purified to apparent homogeneity. The crucial factor leading to the production of active [Glu304]MCADH protein is the expression in E. coli cells at reduced temperature (28 degrees C). Expression in the same system at 37 degrees C results in very low amounts of active mutant protein. Several catalytic and physicochemical parameters of these two proteins have been determined and were compared to those of purified pig kidney MCADH. Although [Glu304]MCADH has approximately the same rate of substrate reduction with dodecanoyl-CoA and the same V(max) as human MCADH with the best substrate for the latter, octanoyl-CoA, the K(m) in the mutant MCADH is fourfold higher, which generates a correspondingly lower catalytic efficiency. Importantly, V(max) obtained using the natural acceptor, electron transfer flavoprotein, is only a third that for human MCADH. The V(max)/K(m) versus chain-length profile of the mutant shows a maximum with dodecanoyl-CoA which differs markedly from that of human MCADH, which has maximal efficiency with octanoyl-CoA. The substrate specificity of the mutant is broader with a less pronounced activity peak resembling long-chain acyl-CoA dehydrogenase. The purified mutant enzyme exhibits a reduced thermal stability compared to human wild-type MCADH. The major difference between the two proteins expressed in E. coli is the more pronounced lability of the K304E mutant in crude extracts, which suggests a higher susceptibility to attack by endogenous proteases. Differences between tetrameric [Glu304]MCADH which survives the first step(s) of purification and corresponding MCADH are minor. The overall differences in properties of [Glu304]MCADH together with its impaired folding and tetramer assembly may contribute to the generation of the abnormalities observed in patients homozygous for the K304E mutation.


RNA Sequencing for Gene Expression Profiles in Peripheral Blood Mononuclear Cells with Ankylosing Spondylitis RNA.
Several previous studies have attempted to investigate the regulatory mechanisms underlying gene expression in ankylosing spondylitis (AS). However, the specific molecular pathways underlying this condition remain unclear. Previous research used next-generation RNA sequencing to identify a series of differentially expressed genes (DEGs) in peripheral blood mononuclear cells (PBMCs) when compared between patients with AS and healthy controls, thus implying that these DEGs may be related to AS. Furthermore, by screening these DEGS, it may be possible to facilitate clinical diagnosis and optimize treatment strategies. In order to test this hypothesis, we recruited 15 patients with AS and 15 healthy controls. We randomly selected five subjects from each group of patients for RNA sequencing analysis. Sequence reads were generated by an Illumina HiSeq2500 platform and mapped on to the human reference genome using HISAT2. We successfully identified 973 significant DEGs (p < 0.05) in PBMCs. When compared with controls, 644 of these genes were upregulated (with a fold change (FC) > 2) in AS patients and 329 were downregulated (FC < 0.5). Our analysis identified numerous genes related to immune response. Gene Ontology (GO) analysis indicated that these DEGs were significantly related to the positive regulation of epidermal growth factor-activated receptor activity, the positive regulation of the ERBB (erb-b2 receptor tyrosine kinase) signaling pathway, the differentiation of trophoblast giant cells, oxygen transport, immune-related pathways, and inflammation-related pathways. The DEGs were also closely related to the TNF and NF-κB signaling pathways. Six DEGs were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Receiver operating characteristic (ROC) curve analysis indicated that IL6 may represent a useful biomarker for diagnosing AS. The development of new biomarkers may help us to elucidate the specific mechanisms involved in the development and progression of AS.


Fighting Ebola with novel spore decontamination technologies for the military.
Recently, global public health organizations such as Doctors without Borders (MSF), the World Health Organization (WHO), Public Health Canada, National Institutes of Health (NIH), and the U.S. government developed and deployed Field Decontamination Kits (FDKs), a novel, lightweight, compact, reusable decontamination technology to sterilize Ebola-contaminated medical devices at remote clinical sites lacking infra-structure in crisis-stricken regions of West Africa (medical waste materials are placed in bags and burned). The basis for effectuating sterilization with FDKs is chlorine dioxide (ClO2) produced from a patented invention developed by researchers at the US Army Natick Soldier RD&E Center (NSRDEC) and commercialized as a dry mixed-chemical for bacterial spore decontamination. In fact, the NSRDEC research scientists developed an ensemble of ClO2 technologies designed for different applications in decontaminating fresh produce; food contact and handling surfaces; personal protective equipment; textiles used in clothing, uniforms, tents, and shelters; graywater recycling; airplanes; surgical instruments; and hard surfaces in latrines, laundries, and deployable medical facilities. These examples demonstrate the far-reaching impact, adaptability, and versatility of these innovative technologies. We present herein the unique attributes of NSRDEC's novel decontamination technologies and a Case Study of the development of FDKs that were deployed in West Africa by international public health organizations to sterilize Ebola-contaminated medical equipment. FDKs use bacterial spores as indicators of sterility. We review the properties and structures of spores and the mechanisms of bacterial spore inactivation by ClO2. We also review mechanisms of bacterial spore inactivation by novel, emerging, and established non-thermal technologies for food preservation, such as high pressure processing, irradiation, cold plasma, and chemical sanitizers, using an array of Bacillus subtilis mutants to probe mechanisms of spore germination and inactivation. We employ techniques of high-resolution atomic force microscopy and phase contrast microscopy to examine the effects of γ-irradiation on bacterial spores of Bacillus anthracis, Bacillus thuringiensis, and Bacillus atrophaeus spp. and of ClO2 on B. subtilis spores, and present in detail assays using spore bio-indicators to ensure sterility when decontaminating with ClO2.


Clinical profiles, endoscopic and laboratory features and associated factors in patients with autoimmune gastritis.
Autoimmune gastritis (AIG) may predispose to gastric carcinoid tumors or adenocarcinomas and may also cause unexplained iron and/or vitamin B(12) deficiency. The aims of this study were to explore clinical manifestations, endoscopic findings and laboratory features of patients with AIG. 109 patients with AIG were enrolled into the study. In addition to demographic and clinical data, gastric lesions, serum gastrin, vitamin B(12), antiparietal cell antibody (APA), current Helicobacter pylori status, and anti-H. pylori IgG were also investigated. The mean age of the patients was 53.06 ± 12.7 years (range 24-81; 72 (66.1%) women). The most common main presenting symptom was abdominal symptoms in 51 patients, consultation for iron and/or vitamin B(12) deficiency in 36, and non-specific symptoms including intermittent diarrhea in 15 patients. Endoscopic lesions were detected in 17 patients, hyperplastic polyps in 8, gastric carcinoid tumor in 4, fundic gland polyps in 3, and adenomatous polyps in 2 patients. H. pylori was negative in all patients in biopsy specimens; however, anti-H. pylori IgG was positive in 30 (27.5%) patients. 91 patients (83.4%) were positive for APA. In patients with AIG, the main symptoms prompted for clinical investigation were: abdominal symptoms, iron/B(12) deficiency and non-specific symptoms. 20% of patients with AIG had various gastric lesions including type I gastric carcinoids. None of the patients were positive for H. pylori by means of invasive tests; however, anti-H. pylori IgG was found in 27.5% of patients. Patients referring with non-specific abdominal symptoms such as bloating, diarrhea and iron/B(12) deficiency should be investigated for the presence of AIG.


Metabolic rivalry: circadian homeostasis and tumorigenesis.
Circadian rhythms govern a large array of physiological and metabolic functions. Perturbations of the daily cycle have been linked to elevated risk of developing cancer as well as poor prognosis in patients with cancer. Also, expression of core clock genes or proteins is remarkably attenuated particularly in tumours of a higher stage or that are more aggressive, possibly linking the circadian clock to cellular differentiation. Emerging evidence indicates that metabolic control by the circadian clock underpins specific hallmarks of cancer metabolism. Indeed, to support cell proliferation and biomass production, the clock may direct metabolic processes of cancer cells in concert with non-clock transcription factors to control how nutrients and metabolites are utilized in a time-specific manner. We hypothesize that the metabolic switch between differentiation or stemness of cancer may be coupled to the molecular clockwork. Moreover, circadian rhythms of host organisms appear to dictate tumour growth and proliferation. This Review outlines recent discoveries of the interplay between circadian rhythms, proliferative metabolism and cancer, highlighting potential opportunities in the development of future therapeutic strategies.


Morning breathing exercises prolong lifespan by improving hyperventilation in people living with respiratory cancer.
Disturbance of oxygen-carbon dioxide homeostasis has an impact on cancer. Little is known about the effect of breath training on cancer patients. Here we report our 10-year experience with morning breathing exercises (MBE) in peer-support programs for cancer survivors.We performed a cohort study to investigate long-term surviving patients with lung cancer (LC) and nasopharyngeal cancer (NPC) who practiced MBE on a daily basis. End-tidal breath holding time (ETBHT) after MBE was measured to reflect improvement in alveolar O2 pressure and alveolar CO2 pressure capacity.Patients (female, 57) with a diagnosis of LC (90 patients) and NPC (32 patients) were included. Seventy-six of them were MBE trainees. Average survival years were higher in MBE trainees (9.8 ± 9.5) than nontrainees (3.3 ± 2.8). The 5-year survival rate was 56.6% for MBE trainees and 19.6% for nontrainees (RR = 5.371, 95% CI = 2.271-12.636, P < 0.001). Survival probability of the trainees further increased 17.9-fold for the 10-year survival rate. Compared with the nontrainees, the MBE trainees shows no significant differences in ETBHT (baseline, P = 0.795; 1-2 years, P = 0.301; 3-4 years, P = 0.059) at baseline and within the first 4 years. From the 5th year onwards, significant improvements were observed in ETBHT, aCO2%, PaCO2, and PaO2 (P = 0.028). In total, 18 trainees (40.9%) and 20 nontrainees (74.1%) developed new metastasis (RR = 0.315, 95% CI = 0.108-0.919, P = 0.031).MBE might benefit for the long-term survival in patients with LC and NPC due to improvement in hyperventilation.


Selective stimulation of small cell lung cancer clonal growth by bombesin and gastrin-releasing peptide.
Human small cell lung cancers (SCLC) produce and secrete the regulatory peptide bombesin (BN) or its mammalian counterpart gastrin-releasing peptide (GRP). In addition, several SCLC tumor lines have been shown to express high affinity receptors for BN/GRP. On the basis of these findings, we investigated the effect of exogenously added BN and GRP on the soft agarose colony growth of a panel of human cell lines. In serum-free defined medium, colony formation of 9 of 10 SCLC cell lines was stimulated up to 150-fold by BN or GRP, with peak colony stimulation observed at 50 nM BN. In contrast, no stimulatory effect of BN was observed on nine non-SCLC cell lines. Although no stimulation of colony growth by BN was seen in serum-supplemented medium, addition of BN to the serum-free medium increased cloning efficiency to that achieved by serum in most of the SCLC cell lines. GRP 1-27, the active mammalian analogue of Bn, stimulated colony growth of SCLC cells similar to the manner of BN, while the physiologically inactive BN analogue, des-Leu (13)-Met (14)-BN, had no effect on colony growth. No correlation was observed in SCLC cell lines between the response of these cells to exogenous BN and the amount of cellular BN/GRP produced or the presence of BN receptors. These data suggest that BN/GRP may in some instances function as an autocrine growth factor for SCLC and indicate new ways for modulating SCLC growth in patients with this tumor.


Evidence-based management of incidental focal uptake of fluorodeoxyglucose on PET-CT.
Focal incidental uptake, with or without CT abnormalities, is a common finding on fluorodeoxyglucose PET/CT and evidence-based management for this type of uptake is lacking. This article reviews the evidence on focal incidental uptake including the incidence of malignancy, differential diagnosis and imaging criteria which can be used to further characterize it. The article focusses on PET rather than CT criteria. The strength of the evidence base is highly variable ranging from systematic reviews and meta-analyses to a virtual absence of evidence. Caution needs to be used when using standardized uptake values (SUVs) reported in other studies due to interpatient and institution observed variation in SUVs. There is sufficient evidence to permit specific suggestions on how to interpret the foci and recommend further management in the: pituitary (investigate when SUVmax >4.1), thyroid (investigate all), breast (investigate all), lung parenchyma (if focus of fluorodeoxyglucose without a CT nodule, no further investigations), colon (investigate all foci with SUVmax >5.9, urgently if SUVmax >11.4), adrenals (criteria depend on if patient has cancer) and prostate gland (investigate in males aged >50 years or >40 years if peripheral uptake or patient has other risk factors). There is some evidence to guide further management for the parotid gland, naso-orophaynx, oesophagus, pancreas, uterus and ovaries. There is insufficient evidence to guide management for the liver, spleen, kidneys, gallbladder, testis and bone, for these organs patient characteristics and other guidelines will likely be of more use in determining further management.


Esmolol inhibits inflammation and apoptosis in the intestinal tissue via the overexpression of NF-κB-p65 in the early stage sepsis rats.
Accumulating evidence reveals esmolol could protect the gut mucosa through the regulation of immune response and inflammation in patients with sepsis. However, its underlying mechanism is not fully understood. Diamine oxidase (DAO), intestinal fatty acid-binding protein (I-FABP), interleukin (IL)-6, and IL-10 in the plasma of rats were detected by ELISA assay. Western blotting was utilized to measure the expression levels of NF-kappa B-p65, Bcl-2, and cleaved caspase-3 in the intestinal tissues. The survival analysis was performed in each group. The plasma levels of DAO and IL-10 levels were increased, whereas that of I-FABP and IL-6 were decreased in the sepsis rats after esmolol treatment, indicating that after the esmolol treatment, the intestinal inflammation and damages were remarkably reduced as compared to those in the normal saline treated sepsis rats. NF-κB-p65 and Bcl-2 were highly expressed, but cleaved caspase-3 showed lower expression in the esmolol treated groups. However, at the same time, we observed contrasting results in the normal saline treated group. Western blotting data indicated that the esmolol treatment inhibited the inflammation and apoptosis in the intestinal tissue due to the overexpression of NF-κB-p65 in the celiac sepsis rats. The survival analysis results indicate that the esmolol infusion should be used in the early stages sepsis rats. Esmolol can suppress inflammation and apoptosis in the intestinal tissue via the overexpression of NF-kappa B-p65 in the early stage sepsis rats. kappa BEarly-stage use of esmolol might be an ideal treatment method for sepsis.


Xenopus fibrillin is expressed in the organizer and is the earliest component of matrix at the developing notochord-somite boundary.
We identify a Xenopus fibrillin homolog (XF), and show that its earliest developmental expression is in presumptive dorsal mesoderm at gastrulation, and that XF expression is regulated by mesoderm-inducing factors in animal cap assays. XF protein is also first detected in presumptive mesoderm, but is concentrated specifically into extracellular-matrix structures that begin to develop de novo by mid-gastrulation at both of the bilateral presumptive notochord-somite boundaries. Later in embryogenesis, XF protein is localized to the extracellular matrix at tissue boundaries, where it is found surrounding the notochord, the somites, and the neural tube, as well as under the epidermis. This pattern of protein deposition combines to give the appearance of an "embryonic skeleton," suggesting that one role for XF is to serve as a mechanical element in the embryo prior to bone deposition.


Osteoporosis--an update.
Osteoporosis and subsequent fractures is a rapidly growing major health problem in many parts of the world. Even if prevalence and incidence are high in Europe, rate of increase is even higher in Asia. The majority of fractures occur in women underlining the importance of ovarian deficiency. Genetic factors and lifestyle greatly influence the incidence and prevalence both in men and women and the importance of estrogen deficiency at ages over 85, where a large number of fractures actually occur, may well be questioned. Among several risk factors the amount of bone mass remains crucial. Several techniques adequate in accuracy and precision exist to determine bone mass. The costs for these measurements are not negligible. Several pharmacologic regimens look promising for the prevention of osteoporosis and indeed for treatment of established osteoporosis. However, long-term data and clinical experience are warranted prior to establishing these pharmacological tools in the field of osteoporosis. For HRT and ERT such data are available. ERT and indeed HRT may halve the risk of osteoporotic fractures in women at least in current users. The effect at least on bone mass wears off after discontinuation of treatment but is never totally lost. In the interest of health care costs it is suggested that HRT could be started 15-20 years after the menopause and still provide protection for subsequent fractures.


In silico local structure approach: a case study on outer membrane proteins.
The detection of Outer Membrane Proteins (OMP) in whole genomes is an actual question, their sequence characteristics have thus been intensively studied. This class of protein displays a common beta-barrel architecture, formed by adjacent antiparallel strands. However, due to the lack of available structures, few structural studies have been made on this class of proteins. Here we propose a novel OMP local structure investigation, based on a structural alphabet approach, i.e., the decomposition of 3D structures using a library of four-residue protein fragments. The optimal decomposition of structures using hidden Markov model results in a specific structural alphabet of 20 fragments, six of them dedicated to the decomposition of beta-strands. This optimal alphabet, called SA20-OMP, is analyzed in details, in terms of local structures and transitions between fragments. It highlights a particular and strong organization of beta-strands as series of regular canonical structural fragments. The comparison with alphabets learned on globular structures indicates that the internal organization of OMP structures is more constrained than in globular structures. The analysis of OMP structures using SA20-OMP reveals some recurrent structural patterns. The preferred location of fragments in the distinct regions of the membrane is investigated. The study of pairwise specificity of fragments reveals that some contacts between structural fragments in beta-sheets are clearly favored whereas others are avoided. This contact specificity is stronger in OMP than in globular structures. Moreover, SA20-OMP also captured sequential information. This can be integrated in a scoring function for structural model ranking with very promising results.


Uptake of Cyclic AMP by Natural Populations of Marine Bacteria.
The major objective of this study was to describe the mechanism(s) of cyclic AMP uptake by natural populations of marine bacteria. A second objective was to determine whether this uptake could contribute to the intracellular regulatory pool of cyclic AMP. Using high-specific-activity P-labeled cyclic AMP, we found several high-affinity uptake systems. The highest-affinity system had a half-saturation constant of <10 pM. This system was extremely specific for cyclic nucleotides, particularly cyclic AMP. It appeared to meet the criteria for active transport. Uptake of cyclic AMP over a wide concentration range (up to 2 muM) showed multiphasic kinetics, with half-saturation constants of 1 nM and greater. These lower-affinity systems were much less specific for cyclic nucleotides. Although much of the labeled cyclic AMP taken up by the high-affinity systems was metabolized, some remained as intact cyclic AMP within the cells during 1 h of incubation. This suggests that at least some of the bacteria use cyclic AMP dissolved in seawater to augment their intracellular pools.


Establishment of pemetrexed-resistant non-small cell lung cancer cell lines.
Pemetrexed (PEM), a multitargeted antifolate with manageable toxicity, is active against non-squamous non-small cell lung cancer; however, most patients eventually acquire resistance to PEM. To elucidate the resistant mechanism, we established PEM-resistant lung adenocarcinoma cell lines. Two parental cell lines, PC-9 and A549, were treated with step-wise increasing concentrations of PEM. Growth inhibition was determined by the 3-[4,5-dimethyl-thizol-2-yl]-2,5-diphenyltetrazolium bromide assay. Expression of the genes encoding thymidylate synthase (TS), dihydrofolate reductase (DHFR), and glycinamide ribonucleotide formyltransferase (GARFT) was analyzed by quantitative real-time reverse transcriptase polymerase chain reaction. The four PC-9 sublines were more resistant than the PC-9 cell line to PEM (2.2-, 2.9-, 8.4-, and 14.3-fold, respectively). The four A549 sublines also showed more resistance to PEM (7.8-, 9.6-, 42.3-, and 42.4-fold, respectively) than the parent cell line. All resistant sublines showed cross-resistance to cisplatin, but not to docetaxel, vinorelbine, 5-fluorouracil, or the active metabolite of irinotecan, SN-38. All PEM-resistant sublines expressed more TS than the parental cells, by polymerase chain reaction and Western blotting. DHFR was significantly increased in the four PEM-resistant A549 sublines. GARFT did not correlate with resistance to PEM. In summary, PEM-resistant cells remained sensitive to docetaxel, vinorelbine, 5-fluorouracil, and irinotecan. TS expression appeared to be associated with resistance to PEM.


OsFIE2 plays an essential role in the regulation of rice vegetative and reproductive development.
Polycomb group (PcG) proteins are gene repressors that help to maintain cellular identity during development via chromatin remodeling. Fertilization-independent endosperm (FIE), a member of the PcG complex, operates extensively in plant development, but its role in rice has not been fully investigated to date. We report the isolation and characterization of a PcG member in rice, which was designated OsFIE2 for Oryza sativa Fertilization-Independent Endosperm 2. OsFIE2 is a single-copy gene in the rice genome and shows a universal expression pattern. The OsFIE2 RNAi lines displayed pleiotropic phenotypes in vegetative and reproductive organ generation. In unfertilized lines, endosperm formation could be triggered without embryo formation, which indicates that FIE is indeed involved in the suppression of autonomous endosperm development in rice. Furthermore, lateral root generation was promoted early in the roots of OsFIE2 RNAi lines, whereas the primary root was premature and highly differentiated. As the root tip stem cell differentiated, QHB, the gene required for stem cell maintenance in the quiescent center, was down-regulated. Our data suggest that the OsFIE2-PcG complex is vital for rice reproduction and endosperm formation. Its role in stem cell maintenance suggests that the gene is functionally conserved in plants as well as animals.


Extraintestinal Manifestations of Inflammatory Bowel Disease: Current Concepts, Treatment, and Implications for Disease Management.
Inflammatory bowel diseases (IBDs) are systemic diseases that manifest not only in the gut and gastrointestinal tract, but also in the extraintestinal organs in many patients. The quality of life for patients with IBD can be substantially affected by these extraintestinal manifestations (EIMs). It is important to have knowledge of the prevalence, pathophysiology, and clinical presentation of EIMs in order to adapt therapeutic options to cover all aspects of IBD. EIMs can occur in up to 24% of patients with IBD before the onset of intestinal symptoms, and need to be recognized to initiate appropriate diagnostic procedures. EIMs most frequently affect joints, skin, or eyes, but can also affect other organs, such as the liver, lung, and pancreas. It is a frequent misconception that a successful therapy of the intestinal inflammation will be sufficient to treat EIMs satisfactorily in most patients with IBD. In general, peripheral arthritis, oral aphthous ulcers, episcleritis, or erythema nodosum can be associated with active intestinal inflammation and can improve on standard treatment of the intestinal inflammation. However, anterior uveitis, ankylosing spondylitis, and primary sclerosing cholangitis usually occur independent of disease flares. This review provides a comprehensive overview of epidemiology, pathophysiology, clinical presentation, and treatment of EIMs in IBD.


Development of Safe and Non-Self-Immunogenic Mucosal Adjuvant by Recombinant Fusion of Cholera Toxin A1 Subunit with Protein Transduction Domain.
Potential use of cholera toxin (CT) as a mucosal vaccine adjuvant has been documented in a variety of animal models. However, native CT is highly toxic to be used as a mucosal adjuvant in humans. Here, we demonstrate a new approach to generate a mucosal adjuvant by replacing the B subunit of CT with HIV-1 Tat protein transduction domain (PTD), which efficiently delivers fusion proteins into the cell cytoplasm by unspecific binding to cell surface. We compared the adjuvanticity and toxicity of Tat PTD-CTA1-Tat PTD (TCTA1T) with those of CT. Our results indicate that intranasal (i.n.) delivery of ovalbumin (OVA) with TCTA1T significantly augments the OVA-specific systemic and mucosal antibody responses to levels comparable to those seen with CT adjuvant. Moreover, in vivo cytotoxic T lymphocyte activity elicited by TCTA1T was significantly higher than that elicited by a mutant TCTA1T (TmCTA1T) lacking ADP-ribosyltransferase function. In addition, coadministration of influenza M2 protein with TCTA1T conferred near complete protection against lethal influenza virus challenge. Importantly, TCTA1T, in contrast to CT, did not induce serum IgG antibody responses to itself and was shown to be nontoxic. These results suggest that TCTA1T may be a safe and effective adjuvant when given by mucosal routes.


Pathogen proteins eliciting antibodies do not share epitopes with host proteins: a bioinformatics approach.
The best way to prevent diseases caused by pathogens is by the use of vaccines. The advent of genomics enables genome-wide searches of new vaccine candidates, called reverse vaccinology. The most common strategy to apply reverse vaccinology is by designing subunit recombinant vaccines, which usually generate an humoral immune response due to B-cell epitopes in proteins. A major problem for this strategy is the identification of protective immunogenic proteins from the surfome of the pathogen. Epitope mimicry may lead to auto-immune phenomena related to several human diseases. A sequence-based computational analysis has been carried out applying the BLASTP algorithm. Therefore, two huge databases have been created, one with the most complete and current linear B-cell epitopes, and the other one with the surface-protein sequences of the main human respiratory bacterial pathogens. We found that none of the 7353 linear B-cell epitopes analysed shares any sequence identity region with human proteins capable of generating antibodies, and that only 1% of the 2175 exposed proteins analysed contain a stretch of shared sequence with the human proteome. These findings suggest the existence of a mechanism to avoid autoimmunity. We also propose a strategy for corroborating or warning about the viability of a protein linear B-cell epitope as a putative vaccine candidate in a reverse vaccinology study; so, epitopes without any sequence identity with human proteins should be very good vaccine candidates, and the other way around.


The Impact of Specialized Corpora for Word Embeddings in Natural Langage Understanding.
Recent studies in the biomedical domain suggest that learning statistical word representations (static or contextualized word embeddings) on large corpora of specialized data improve the results on downstream natural language processing (NLP) tasks. In this paper, we explore the impact of the data source of word representations on a natural language understanding task. We compared embeddings learned with Fasttext (static embedding) and ELMo (contextualized embedding) representations, learned either on the general domain (Wikipedia) or on specialized data (electronic health records, EHR). The best results were obtained with ELMo representations learned on EHR data for the two sub-tasks (+7% and +4% of gain in F1-score). Moreover, ELMo representations were trained with only a fraction of the data used for Fasttext.