Pathophysiology of platelet-vessel interactions.
Thrombosis results from the interaction of a number of processes which are summarized in table I and figure 1. The predominant role of platelets in the genesis of rapid-flow, arterial thrombosis provides a rationale for the clinical trial of inhibitors of platelet functions in the management of several thrombotic disorders.


Presynaptic membrane retrieval and endosome biology: defining molecularly heterogeneous synaptic vesicles.
The release and uptake of neurotransmitters by synaptic vesicles is a tightly controlled process that occurs in response to diverse stimuli at morphologically disparate synapses. To meet these architectural and functional synaptic demands, it follows that there should be diversity in the mechanisms that control their secretion and retrieval and possibly in the composition of synaptic vesicles within the same terminal. Here we pay particular attention to areas where such diversity is generated, such as the variance in exocytosis/endocytosis coupling, SNAREs defining functionally diverse synaptic vesicle populations and the adaptor-dependent sorting machineries capable of generating vesicle diversity. We argue that there are various synaptic vesicle recycling pathways at any given synapse and discuss several lines of evidence that support the role of the endosome in synaptic vesicle recycling.


Hofmeister effects in surface tension of aqueous electrolyte solution.
The surface tension of electrolyte solutions shows marked specific ion effects. We here show an important role for both ionic solvation energies and ionic dispersion potentials in determining this ion specific surface tension of salt solutions. The ion self-free energy changes when an ion moves from bulk solution into the interfacial region, with its decreasing water density profile. We will show that the solvation energies of different ions correlate very well with the surface tension of salt solutions. Inclusion of this distance-dependent self-free energy contribution brings qualitative agreement with experiments and the right Hofmeister series. This is so not only for surface tension changes but also for measured surface potentials. The inclusion of ionic dispersion interaction potentials further improves the agreement with experiments. We discuss how further progress in the theory of the surface tension of salts can be achieved.


[Nucleic acids utilized as the main source of bacterial nutrition].
Secretion of DNases and RNases, respectively, was found in saprophyte bacteria isolated from nature and growing on media containing DNA and RNA. Serratia marcescens and Bacillus subtilis with a high nuclease activity can assimilate RNA and DNA as the main source of nutrition. Ser. marcescens with a nuclease which attacks both DNA and RNA can grow equally well on these acids. Bac. subtilis has a higher activity of RNase and grows better on RNA.


The development of cytochrome b-245 in maturing human macrophages.
Cytochrome b-245, the putative terminal component of the specialized cidal oxidase system of phagocytes, was measured in human monocytes in culture. There was a dramatic synthesis of the cytochrome, which increased by 27.3 +/- 2.0 pmol/day per 10(7) cells. This represents an increase of about 40%/day in the early stages and an overall 7-fold increase after 16 days. The protein content increased 3-fold over the same period, resulting in a doubling of the specific content of the cytochrome b. The newly synthesized cytochrome b was identified as that specifically located in the microbicidal oxidase electron-transport chain, as titration demonstrated that, at day 16 of maturation, 70% of the total membrane cytochrome b had a very low midpoint potential (-260 to -220 mV), characteristic of that found in this oxidase system. This cytochrome distributed with the plasma membrane on analytical subcellular fractionation, and a close relationship was observed between the maturation-induced increase in the concentration of this molecule and the capacity of the cells to produce superoxide.


A comparison of three different concentrations of levobupivacaine for caudal block in children.
We investigated three different concentrations of levobupivacaine (0.125%, 0.20%, and 0.25%; n = 20 in each group) for caudal blockade in a prospective, randomized, observer-blinded fashion in children (1-7 yr) undergoing subumbilical surgery. The duration of postoperative analgesia was assessed as the time to first administration of supplemental analgesia (based on a Childrens and Infants Postoperative Pain Scale score of >or=4), and the degree of immediate postoperative motor blockade was determined by use of a 3-point scale. A dose-response relationship was observed both with regard to median duration of postoperative analgesia (0.125%, 60 min; 0.20%, 118 min; 0.25%, 158 min) and the number of patients with evidence of early postoperative motor blockade (0.125%, 0; 0.20%, 4; 0.25%, 8). The 0.125% concentration was associated with significantly less early motor blockade (P = 0.003) but was found to result in a significantly shorter duration of postoperative analgesia (P < 0.05). Based on these results, the use of 0.20% levobupivacaine might represent the best clinical option if a plain levobupivacaine solution is to be used for caudal blockade in children. The use of 0.125% levobupivacaine for caudal blockade (1 mL/kg) in children (1-7 yr) was associated with less early postoperative motor blockade but a shorter duration of postoperative analgesia compared with 0.20% and 0.25% solutions.


A meta-analysis of somatic mutations from next generation sequencing of 241 melanomas: a road map for the study of genes with potential clinical relevance.
Next generation sequencing (NGS) has been used to characterize the overall genomic landscape of melanomas. Here, we systematically examined mutations from recently published melanoma NGS data involving 241 paired tumor-normal samples to identify potentially clinically relevant mutations. Melanomas were characterized according to an in-house clinical assay that identifies well-known specific recurrent mutations in five driver genes: BRAF (affecting V600), NRAS (G12, G13, and Q61), KIT (W557, V559, L576, K642, and D816), GNAQ (Q209), and GNA11 (Q209). Tumors with none of these mutations are termed "pan negative." We then mined the driver mutation-positive and pan-negative melanoma NGS data for mutations in 632 cancer genes that could influence existing or emerging targeted therapies. First, we uncovered several genes whose mutations were more likely associated with BRAF- or NRAS-driven melanomas, including TP53 and COL1A1 with BRAF, and PPP6C, KALRN, PIK3R4, TRPM6, GUCY2C, and PRKAA2 with NRAS. Second, we found that the 69 "pan-negative" melanoma genomes harbored alternate infrequent mutations in the five known driver genes along with many mutations in genes encoding guanine nucleotide binding protein α-subunits. Third, we identified 12 significantly mutated genes in "pan-negative" samples (ALK, STK31, DGKI, RAC1, EPHA4, ADAMTS18, EPHA7, ERBB4, TAF1L, NF1, SYK, and KDR), including five genes (RAC1, ADAMTS18, EPHA7, TAF1L, and NF1) with a recurrent mutation in at least two "pan-negative" tumor samples. This meta-analysis provides a road map for the study of additional potentially actionable genes in both driver mutation-positive and pan-negative melanomas.


In vivo assessment of enhanced topical delivery of terbinafine to human stratum corneum.
The objective of this study was to evaluate, using attenuated total reflectance Fourier transform infrared spectroscopy, the stratum corneum (SC) bioavailability of terbinafine (TBF) following topical treatment with four different formulations. Four skin sites on the ventral forearms of five healthy volunteers were treated for 2 h using one of four formulations based on a vehicle consisting of 50% ethanol and 50% isopropyl myristate. Three of these formulations included a percutaneous penetration enhancer: either 5% oleic acid, 10% 2-pyrrolidone or 1% urea. The SC concentration profile of TBF was measured by repeated infrared spectroscopic measurements while sequentially stripping off the layers of this barrier membrane with adhesive tape. This method was validated by HPLC analysis of TBF extracted from the stripped tapes. Transepidermal water loss (TEWL) measurements were also performed, to permit facile estimation of SC thickness. The SC concentration profiles of TBF were fitted to the appropriate solution of Fick's second law of diffusion, thereby allowing determination of the characteristic diffusion and partitioning parameters of the permeating drug. This analysis enabled the efficacies of the different formulations tested to be compared to the no-enhancer control. While it was found that the formulation containing 5% oleic acid significantly enhanced the SC availability of TBF, the other formulations did not improve the apparent drug delivery. A facile and minimally invasive methodology to evaluate an important aspect of topical drug bioavailability has been described. The analytical methods used (infrared spectroscopy and HPLC) allow estimates of both relative and absolute drug bioavailability in the SC and may be useful, therefore, in the critical determination of bioequivalence between topical formulations.


Single DNA sequence common to all chlamydial species employed for PCR detection of these organisms.
Chlamydial infection is responsible for a wide spectrum of diseases of the eye, genitourinary tract, and lung. This group of organisms is also implicated in the pathogenesis of coronary artery disease as well as arthritis. Since cross-species infection is widely reported (though probably underestimated), it is an advantage to have a rapid and reliable method to detect all forms of chlamydiae in patient samples. We have identified a 160/163-bp DNA fragment in Chlamydia which is highly conserved in all chlamydial species. A polymerase chain reaction method based on this sequence has been developed to detect, in clinical samples, chlamydiae which have been shown to be positive by fluorescent-staining immunoassay; this method can be utilized in combination with restriction endonuclease cleavage to identify individual chlamydial species. Thus we have developed a sensitive and rapid detection method and have used it on samples from patients with respiratory and genital infections.


Decreased expression of the ARID1A gene is associated with poor prognosis in primary gastric cancer.
The ARID1A gene encodes adenine-thymine (AT)-rich interactive domain-containing protein 1A, which participates in chromatin remodeling. ARID1A has been showed to function as a tumor suppressor in various cancer types. In the current study, we investigated the expression and prognosis value of ARID1A in primary gastric cancer. Meanwhile, the biological role of ARID1A was further investigated using cell model in vitro. To investigate the role of ARID1A gene in primary gastric cancer pathogenesis, real-time quantitative PCR and western blotting were used to examine the ARID1A expression in paired cancerous and noncancerous tissues. Results revealed decreased ARID1A mRNA (P = 0.0029) and protein (P = 0.0015) expression in most tumor-bearing tissues compared with the matched adjacent non-tumor tissues, and in gastric cancer cell lines. To further investigate the clinicopathological and prognostic roles of ARID1A expression, we performed immunohistochemical analyses of the 224 paraffin-embedded gastric cancer tissue blocks. Data revealed that the loss of ARID1A expression was significantly correlated with T stage (P = 0.001) and grade (P = 0.006). Consistent with these results, we found that loss of ARID1A expression was significantly correlated with poor survival in gastric cancer patients (P = 0.003). Cox regression analyses showed that ARID1A expression was an independent predictor of overall survival (P = 0.029). Furthermore, the functions of ARID1A in the proliferation and colony formation of gastric cell lines were analyzed by transfecting cells with full-length ARID1A expression vector or siRNA targeting ARID1A. Restoring ARID1A expression in gastric cancer cells significantly inhibited cell proliferation and colony formation. Silencing ARID1A expression in gastric epithelial cell line significantly enhanced cell growth rate. Our data suggest that ARID1A may play an important role in gastric cancer and may serve as a valuable prognostic marker and potential target for gene therapy in the treatment of gastric cancer.


Co-expression of P2X1 and P2X5 receptor subunits reveals a novel ATP-gated ion channel.
P2X receptors are a family of ion channels gated by extracellular ATP. Each member of the family can form functional homomeric channels, but only P2X2 and P2X3 have been shown to combine to form a unique heteromeric channel. Data from in situ hybridization studies suggest that P2X1 and P2X5 may also co-assemble. In this study, we tested this hypothesis by expressing recombinant P2X1 and P2X5 receptor subunits either individually or together in human embryonic kidney 293 cells. In cells expressing the homomeric P2X1 receptor, 30 microM alpha,beta-methylene ATP (alpha,beta-me-ATP) evoked robust currents that completely desensitized in less than 1 sec, whereas alpha,beta-me-ATP failed to evoke current in cells expressing the homomeric P2X5 receptor. By contrast, alpha, beta-me-ATP evoked biphasic currents with a pronounced nondesensitizing plateau phase in cells that co-expressed both subunits. Further, the EC50 for alpha,beta-me-ATP was greater in cells expressing both P2X1 and P2X5 than in cells expressing P2X1 alone (5 and 1.6 microM, respectively). Heteromeric assembly was confirmed using a co-immunoprecipitation assay of epitope-tagged P2X1 and P2X5 subunits. In summary, this study provides biochemical and functional evidence of a novel channel formed by P2X subunit heteropolymerization. This finding suggests that heteromeric subunit assembly constitutes an important mechanism for generating functional diversity of ATP-mediated responses.


Deciphering complex patterns of class-I HLA-peptide cross-reactivity via hierarchical grouping.
T-cell responses in humans are initiated by the binding of a peptide antigen to a human leukocyte antigen (HLA) molecule. The peptide-HLA complex then recruits an appropriate T cell, leading to cell-mediated immunity. More than 2000 HLA class-I alleles are known in humans, and they vary only in their peptide-binding grooves. The polymorphism they exhibit enables them to bind a wide range of peptide antigens from diverse sources. HLA molecules and peptides present a complex molecular recognition pattern, as many peptides bind to a given allele and a given peptide can be recognized by many alleles. A powerful grouping scheme that not only provides an insightful classification, but is also capable of dissecting the physicochemical basis of recognition specificity is necessary to address this complexity. We present a hierarchical classification of 2010 class-I alleles by using a systematic divisive clustering method. All-pair distances of alleles were obtained by comparing binding pockets in the structural models. By varying the similarity thresholds, a multilevel classification was obtained, with 7 supergroups, each further subclassifying to yield 72 groups. An independent clustering performed based only on similarities in their epitope pools correlated highly with pocket-based clustering. Physicochemical feature combinations that best explain the basis of clustering are identified. Mutual information calculated for the set of peptide ligands enables identification of binding site residues contributing to peptide specificity. The grouping of HLA molecules achieved here will be useful for rational vaccine design, understanding disease susceptibilities and predicting risk of organ transplants.


Evidence for a form of adrenergic response to stress in the mollusc Crassostrea gigas.
Catecholamines and pro-opiomelanocortin (POMC)-derived peptides, some of the central regulators of the stress-response systems of vertebrates, are also present in invertebrates. However, studies are needed to determine how these hormones participate in the organisation of neuroendocrine stress-response axes in invertebrates. Our present work provides evidence for the presence of an adrenergic stress-response system in the oyster Crassostrea gigas. Noradrenaline and dopamine are released into the circulation in response to stress. Storage and release of these hormones take place in neurosecretory cells presenting morphological and biochemical similarities with vertebrate chromaffin cells. Both in vivo and in vitro experiments showed that applications of the neurotransmitters acetylcholine or carbachol caused no significant release of noradrenaline or dopamine. Moreover, the nicotinic antagonists hexamethonium and &agr; -bungarotoxin and the muscarinic antagonist atropine caused no significant inhibition of catecholamine release in stressed oysters. Adrenocorticotropic hormone (ACTH) induced a significant release of noradrenaline, but the release of dopamine in response to ACTH was not significant. These results suggest that, unlike that of vertebrates, the adrenergic stress-response system of oysters is not under the control of acetylcholine and that other factors, such as the neuropeptide ACTH, might control this system.


Dynamics of slow-moving landslides from permanent scatterer analysis.
High-resolution interferometric synthetic aperture radar (InSAR) permanent scatterer data allow us to resolve the rates and variations in the rates of slow-moving landslides. Satellite-to-ground distances (range changes) on landslides increase at rates of 5 to 7 millimeters per year, indicating average downslope sliding velocities from 27 to 38 millimeters per year. Time-series analysis shows that displacement occurs mainly during the high-precipitation season; during the 1997-1998 El Niño event, rates of range change increased to as much as 11 millimeters per year. The observed nonlinear relationship of creep and precipitation rates suggests that increased pore fluid pressures within the shallow subsurface may initiate and accelerate these features. Changes in the slope of a hill resulting from increases in the pore pressure and lithostatic stress gradients may then lead to landslides.


Antithrombotic efficacy of low-molecular-weight heparin in deep arterial injury.
Low-molecular-weight heparin subfractions more specifically inhibit factor Xa than thrombin, and they may have advantages over unfractionated heparin in arterial thrombosis. The antithrombotic efficacy of four dosages of a low-molecular-weight heparin (CY216 at 100, 200, 400, or 500 Institute Choay units/kg) was compared with unfractionated calcium heparin (100 US Pharmacopeia units/kg) and placebo during deep arterial injury produced by balloon dilatation of the carotid artery in the pig. The acute thrombotic end points were 111In-labeled platelet and 125I-labeled fibrinogen/fibrin deposition and macroscopic mural thrombosis; these were related to the anti-factor Xa and antithrombin effects of the heparin preparations. Platelet deposition in segments with deep arterial injury was 42 +/- 28, 22 +/- 5, 29 +/- 12, 9 +/- 2, 9 +/- 2, and 11 +/- 3 x 10(6)/cm2 (mean +/- SEM) for pigs treated with placebo, with 100, 200, 400, and 500 units/kg CY216, and with 100 units/kg unfractionated heparin, respectively. Fibrinogen/fibrin deposition was 35 +/- 8, 19 +/- 2, 19 +/- 4, 21 +/- 3, 14 +/- 4, and 12 +/- 3 molecules x 10(12)/cm2, respectively; deposition was significantly reduced in pigs given 100 units/kg unfractionated heparin compared with placebo (p less than 0.05). Mural thrombosis was present in 74%, 45%, 30%, 14%, 5%, and 9% of deeply injured arterial segments, respectively (p = 0.02). Plasma anti-factor Xa activity and prolongation of the activated partial thromboplastin time (aPTT) with 100 units/kg unfractionated heparin were similar to that produced by 200 units/kg and 500 units/kg CY216, respectively. Thus, low-molecular-weight heparin, which predominantly inhibits factor Xa activity, was only moderately effective at reducing platelet thrombus deposition. It was less effective than 100 units/kg unfractionated heparin, except at high dosages, producing similar prolongation of the aPTT and the thrombin time.(ABSTRACT TRUNCATED AT 250 WORDS)


Detection of N-glycolyl-neuraminic acid-containing glycolipids in human skin.
Humans lack the enzyme that produces the sialic acid N-glycolyl neuraminic acid (Neu5Gc), but several lines of evidence have shown that Neu5Gc can be taken up by mammalian food sources and replace the common human sialic acid N-acetyl neuraminic acid (Neu5Ac) in glycans. Cancer tissue has been shown to have increased the presence of Neu5Gc and Neu5Gc-containing glycolipids such as the ganglioside GM3, which have been proposed as tumor-specific antigens for antibody treatment. Here, we show that a previously described antibody against Neu5Gc-GM3 is binding to Neu5GC-containing gangliosides and is strongly staining different cancer tissues. However, we also found a strong intracellular staining of keratinocytes of healthy skin. We confirmed this staining on freshly isolated keratinocytes by flow cytometry and detected Neu5Gc by mass spectrometry. This finding implicates that non-human Neu5Gc can be incorporated into gangliosides in human skin, and this should be taken into consideration when targeting Neu5Gc-containing gangliosides for cancer immunotherapy.


Adding ecosystem function to agent-based land use models.
The objective of this paper is to examine issues in the inclusion of simulations of ecosystem functions in agent-based models of land use decision-making. The reasons for incorporating these simulations include local interests in land fertility and global interests in carbon sequestration. Biogeochemical models are needed in order to calculate such fluxes. The Century model is described with particular attention to the land use choices that it can encompass. When Century is applied to a land use problem the combinatorial choices lead to a potentially unmanageable number of simulation runs. Century is also parameter-intensive. Three ways of including Century output in agent-based models, ranging from separately calculated look-up tables to agents running Century within the simulation, are presented. The latter may be most efficient, but it moves the computing costs to where they are most problematic. Concern for computing costs should not be a roadblock.


Biology of childhood osteogenic sarcoma and potential targets for therapeutic development: meeting summary.
Childhood osteogenic sarcoma (OS) is a rare bone cancer occurring primarily in adolescents. The North American pediatric cooperative groups have performed a series of clinical treatment trials in this disease over the past several decades, and biology studies of tumor tissue have been an important study component. A meeting was held in Bethesda, Maryland on November 29-30, 2001, sponsored by the NIH Office of Rare Diseases, the Children's Oncology Group, and the National Cancer Institute-Cancer Therapy Evaluation Program with the general objectives: (a) to review the current state of knowledge regarding OS biology; (b) to identify, prioritize, and support the development of biology studies of potential clinical relevance in OS; and (c) to discuss the available tissue resources and the appropriate methods for analysis of OS samples for the conduct of biology studies. This report summarizes the information presented and discussed by the meeting participants.


Kidney Injury Causes Accumulation of Renal Sodium That Modulates Renal Lymphatic Dynamics.
Lymphatic vessels are highly responsive to changes in the interstitial environment. Previously, we showed renal lymphatics express the Na-K-2Cl cotransporter. Since interstitial sodium retention is a hallmark of proteinuric injury, we examined whether renal sodium affects NKCC1 expression and the dynamic pumping function of renal lymphatic vessels. Puromycin aminonucleoside (PAN)-injected rats served as a model of proteinuric kidney injury. Sodium 23Na/1H-MRI was used to measure renal sodium and water content in live animals. Renal lymph, which reflects the interstitial composition, was collected, and the sodium analyzed. The contractile dynamics of isolated renal lymphatic vessels were studied in a perfusion chamber. Cultured lymphatic endothelial cells (LECs) were used to assess direct sodium effects on NKCC1. MRI showed elevation in renal sodium and water in PAN. In addition, renal lymph contained higher sodium, although the plasma sodium showed no difference between PAN and controls. High sodium decreased contractility of renal collecting lymphatic vessels. In LECs, high sodium reduced phosphorylated NKCC1 and SPAK, an upstream activating kinase of NKCC1, and eNOS, a downstream effector of lymphatic contractility. The NKCC1 inhibitor furosemide showed a weaker effect on ejection fraction in isolated renal lymphatics of PAN vs controls. High sodium within the renal interstitium following proteinuric injury is associated with impaired renal lymphatic pumping that may, in part, involve the SPAK-NKCC1-eNOS pathway, which may contribute to sodium retention and reduce lymphatic responsiveness to furosemide. We propose that this lymphatic vessel dysfunction is a novel mechanism of impaired interstitial clearance and edema in proteinuric kidney disease.


Genomic Responses of Mouse Synovial Fibroblasts During Tumor Necrosis Factor-Driven Arthritogenesis Greatly Mimic Those in Human Rheumatoid Arthritis.
Aberrant activation of synovial fibroblasts is a key determinant in the pathogenesis of rheumatoid arthritis (RA). The aims of this study were to produce a map of gene expression and epigenetic changes occurring in this cell type during disease progression in the human tumor necrosis factor (TNF)-transgenic model of arthritis and to identify commonalities with human synovial fibroblasts. We used deep sequencing to probe the transcriptome, the methylome, and the chromatin landscape of cultured mouse arthritogenic synovial fibroblasts at 3 stages of disease, as well as synovial fibroblasts stimulated with human TNF. We performed bioinformatics analyses at the gene, pathway, and network levels, compared mouse and human data, and validated selected genes in both species. We found that synovial fibroblast arthritogenicity was reflected in distinct dynamic patterns of transcriptional dysregulation, which was especially enriched in pathways of the innate immune response and mesenchymal differentiation. A functionally representative subset of these changes was associated with methylation, mostly in gene bodies. The arthritogenic state involved highly active promoters, which were marked by histone H3K4 trimethylation. There was significant overlap between the mouse and human data at the level of dysregulated genes and to an even greater extent at the level of pathways. This study is the first systematic examination of the pathogenic changes that occur in mouse synovial fibroblasts during progressive TNF-driven arthritogenesis. Significant correlations with the respective human RA synovial fibroblast data further validate the human TNF-transgenic mouse as a reliable model of the human disease. The resource of data generated in this work may serve as a framework for the discovery of novel pathogenic mechanisms and disease biomarkers.


[Effect of amizil on the carbohydrase activity in intact animals and animals subjected to stress exposure].
Effect of a central cholinolytic drug amizyl on the activity of some digestive enzymes involved in carbohydrate hydrolysis was studied in experiments on control rats and animals exposed to immobilization stress. The experiments showed that amizyl administration modifies the activity of carbohydrases that could affect disintegration and assimilation of carbohydrases. Preliminary administration of the drug was shown to prevent substantially the changes in the enzyme activity occurring during stress situation.


Intersectional Stigma as a Fundamental Cause of Health Disparities: A case study of how drug use stigma intersecting with racism and xenophobia creates health inequities for Black and Hispanic persons who use drugs over time.
Recent evidence points to racial and ethnic disparities in drug-related deaths and health conditions. Informed by stigma, intersectionality, intersectional stigma, and fundamental cause theories, we aimed to explore whether intersectional stigma was a fundamental cause of health. We document key events and policies over time and find that when progress is made new mechanisms emerge that negatively affect health outcomes for Black and Hispanic persons. We then focus on intersectional stigma targeting Black and Hispanic persons who use drugs. We document that when a person, or group of people, occupy multiple stigmatized identities the processes of stigmatization and scapegoating are particularly persistent and pernicious since people and groups can be stigmatized and scapegoated on varying intersections. We propose that an intersectional stigma framework allows for a better understanding of observed patterns over time, thereby providing a better guide for policies and interventions designed to reduce disparities. As a framework, intersectional stigma aims to recognize that when different sources of stigma collide, a new set of circumstances is created for those who reside in the intersection. We conclude that intersectional stigma is a fundamental cause of health inequities and provide policy recommendations aimed at dismantling intersectional stigma processes and mitigating the effects of intersectional stigmas to ultimately promote better health outcomes for Black and Hispanic persons who use drugs.


A developmental increase in the expression of messenger ribonucleic acid encoding a second form of gonadotropin-releasing hormone in the rhesus macaque hypothalamus.
GnRH-I is thought to represent the primary neuroendocrine link between the brain and the reproductive axis. Recently, however, a second molecular form of this decapeptide (GnRH-II) was found to be highly expressed in the brains of humans and nonhuman primates. In this study, in situ hybridization was used to examine the regional expression of GnRH-II messenger ribonucleic acid in the hypothalamus of immature (0.6 yr) and adult (10-15 yr) male and female rhesus macaques (Macaca mulatta). Overall, no sex-related differences were observed. In all of the animals (n = 3 animals/group), intense hybridization of a monkey GnRH-II riboprobe was evident in the paraventricular nucleus and supraoptic nucleus and to a lesser extent in the suprachiasmatic nucleus, but no age- or sex-related differences were apparent. Intense hybridization of the riboprobe also occurred in the mediobasal hypothalamus, and this was markedly greater in the adults than in the immature animals. These data show that the expression of GnRH-II messenger ribonucleic acid increases developmentally in a key neuroendocrine center of the brain. Moreover, because GnRH-II can stimulate LH release in vivo, it is plausible that changes in its gene expression represent an important component of the mechanism by which the hypothalamus controls reproductive function.


Assessing whether an allele can account in part for a linkage signal: the Genotype-IBD Sharing Test (GIST).
To fine map genes, investigators often test for disease-marker association in chromosomal regions with evidence for linkage. Given a marker allele tentatively associated with disease, one would ask if this allele, or one in linkage disequilibrium (LD) with it, could account in part for the observed linkage signal. This question can be addressed by determining if families selected on the basis of the presence of the tentatively associated allele show stronger evidence of linkage as measured by increased allele sharing identical by descent (IBD) by affected family members. However, common selection strategies can be biased for or against linkage in the marker region, even given no disease-marker association. We define unbiased selection schemes and extend the definition to allow weighted selection on the basis of all genotyped family members. For affected-sibship data, we describe three genotype-based weight variables, corresponding to dominant, recessive, and additive models. We then introduce a test for association of a family weight variable with excess IBD sharing. This test allows us to determine if the linkage signal in a region can be attributed in part to the presence of a marker allele, either because of direct involvement in disease etiology or because of LD with a predisposing genetic variant. For samples of 500 affected sib pairs, the tests are powerful in detection of genotype-IBD sharing association, even for disease models with sib relative risk as low as lambda S=1.1, or when evidence for linkage is absent because of sampling variation. This makes our method a new tool for detecting linkage as well as association, especially in regions harboring a candidate gene. We have implemented these methods in the software package GIST (Genotype-IBD Sharing Test).


Two-dimensional IR spectroscopy reveals a hidden Fermi resonance band in the azido stretch spectrum of β-azidoalanine.
Azido stretch modes in a variety of azido-derivatized nonnatural amino acids and nucleotides have been used as a site-specific infrared (IR) probe for monitoring changes in their conformations and local electrostatic environments. The vibrational bands of azide probes are often accompanied by complex line shapes with shoulder peaks, which may arise either from incomplete background subtraction, Fermi resonance, or multiple conformers. The isotope substitution in the infrared probe has thus been introduced to remove Fermi resonances without causing a significant perturbation to the structure. Here, we synthesized and labeled the mid-N atoms of aliphatic azide derivatives with 15N to study the effects of isotope labelling on their vibrational properties. The FT-IR spectra of the aliphatic azide with asymmetric lineshape became a single symmetric band upon isotope substitution, which might be an indication of the removal of the hidden Fermi resonance from the system. We also noticed that the 2D-IR spectrum of unlabeled aliphatic azide has cross-peaks, even though it is not apparently identifiable. The 1D slice spectra obtained from the 2D-IR spectra reveal the existence of a hidden Fermi resonance peak. Furthermore, we show that this weak Fermi resonance does not produce discernible oscillatory beating patterns in the IR pump-probe spectrum, which has been used as evidence of the Fermi resonance. Therefore, we confirm that isotope labelling combined with 2D-IR spectroscopy is the most efficient and incisive way to identify the origin of small shoulder peaks in the linear and nonlinear vibrational spectra of various IR probe molecules.


Quercetin Inhibits Cell Migration and Invasion in Human Osteosarcoma Cells.
Osteosarcoma is a malignant tumor associated with high mortality; however, no effective therapies for the disease have been developed. Several studies have focused on elucidating the pathogenesis of osteosarcoma and have aimed to develop novel therapies for the disease. Quercetin is a vital dietary flavonoid that has been shown to have a variety of anticancer effects, as it induces cell cycle arrest, apoptosis, and differentiation and is involved in cell adhesion, metastasis and angiogenesis. Herein, we aimed to investigate the effects of quercetin on osteosarcoma migration and invasion in vitro and in vivo and to explore the molecular mechanisms underlying its effects on osteosarcoma migration and invasion. Cell viability, cell cycle activity and cell apoptosis were measured using CCK-8 assay and flow cytometry, and cell migration and invasion were evaluated by wound healing and transwell assays, respectively. The mRNA and protein expression levels of several proteins of interest were assessed by real-time quantitative PCR and western blotting, respectively. Moreover, a nude mouse model of human osteosarcoma lung metastasis was established to assess the anti-metastatic effects of quercetin in vivo. We noted no significant differences in cell cycle activity and apoptosis between HOS and MG63 cells and control cells. Treatment with quercetin significantly attenuated cell migration and invasion in HOS and MG63 cells compared with treatment with control medium. Moreover HIF-1α, VEGF, MMP2, and MMP9 mRNA and protein expression levels were significantly downregulated in HOS cells treated with quercetin compared with HOS cells treated with controls. Additionally, treatment with quercetin attenuated metastatic lung tumor formation and growth in the nude mouse model of osteosarcoma compared with treatment with controls. Our findings regarding the inhibitory effects of quercetin on cell migration and invasion suggest that quercetin may have potential as a therapy for human osteosarcoma.


In Silico, in Vitro and in Vivo Study of Substituted Imidazolidinone Sulfonamides as Antibacterial Agents.
New substituted imidazolidinone sulfonamides have been developed using a rational drug design strategy. Predictive QSAR models for the search of new antibacterials were created using the OCHEM platform. Regression models were applied to verify a virtual chemical library of new imidazolidinone derivatives designed to have antibacterial activity. A number of substituted imidazolidinone sulfonamides as effective antibacterial agents were identified by QSAR prediction, synthesized and characterized by spectral and elemental, and tested in vitro. Six studied compounds have shown the highest in vitro antibacterial activity against Gram-negative E. coli and Gram-positive S. aureus multidrug-resistant strains. The in vivo acute toxicity of these imidazolidinone sulfonamides based on the LC50 value ranged from 16.01 to 44.35 mg/L (slightly toxic compounds class). The results of molecular docking suggest that the antibacterial mechanism of the compounds can be associated with the inhibition of post-translational modification processes of bacterial peptides and proteins.


A small RNA promotes siderophore production through transcriptional and metabolic remodeling.
Siderophores are essential factors for iron (Fe) acquisition in bacteria during colonization and infection of eukaryotic hosts, which restrain iron access through iron-binding protein, such as lactoferrin and transferrin. The synthesis of siderophores by Escherichia coli is considered to be fully regulated at the transcriptional level by the Fe-responsive transcriptional repressor Fur. Here we characterized two different pathways that promote the production of the siderophore enterobactin via the action of the small RNA RyhB. First, RyhB is required for normal expression of an important enterobactin biosynthesis polycistron, entCEBAH. Second, RyhB directly represses the translation of cysE, which encodes a serine acetyltransferase that uses serine as a substrate for cysteine biosynthesis. Reduction of CysE activity by RyhB allows serine to be used as building blocks for enterobactin synthesis through the nonribosomal peptide synthesis pathway. Thus, RyhB plays an essential role in siderophore production and may modulate bacterial virulence through optimization of siderophore production.


[Exposure of orbital implants: causes and treatment].
The paper provides the results of a 15-year following-up of 91 patients with exposed orbital implants made from synthetic and biological materials. Surgery was made in the patients treated at different health care facilities. The authors analyze the causes of implant exposure in different periods. Early after surgery the most common causes of implant exposure are surgical equipment failure (inadequate implant coating with adjacent tissues), a too large implant arching the tissues, and mismatch of an artificial eye. In the late period, the more common cause is secondary infection or the toxic properties ofa material itself, which cause a persistent chronic inflammatory reaction. Moreover, the authors consider the mechanism of implant exposure and rejection depending on the material of an implant. The paper describes medical and surgical treatments in patients of this group.


Post-Stroke Resting-State EEG Connectivity: A Longitudinal Neuro-Rehabilitation Study.
Stroke is a leading cause of permanent disability worldwide. Even after adequate treatment, the majority of patients do not recover fully, making them dependent on others for carrying out Activities of Daily Living (ADL). An improved understanding of the underlying mechanism of plasticity will help us in customizing the translational approach for learning and rehabilitation following a stroke. For this study, a 2-minute resting state EEG data were recorded at 5 time-points for 3-months after stroke onset. Directed Transfer Function (DTF) was used to study neural reorganization for 3 months. DTF for different brain regions and sub-bands was correlated with FMA. The information flow was studied for different brain regions as well as Affected Region (AR). Occipital region showed good correlation (r = 0.45 to 0.47) with FMA. Contra-lesional and ipsi-lesional regions trajectories complement each other during acute and sub-acute phase. The information outflow vs inflow imbalance of AR was restored by the end of 3 months. DTF can be used as biomarker for studying neuroplasticity. Occipital, temporal and motor cortex regions play an important role during neuro-rehabilitation. The information about different regions during rehabilitation will help us in designing subject-specific interventions for better recovery.


Results of carotid endarterectomy in patients with contralateral internal carotid artery occlusion from the Mid-America Vascular Study Group and the Society for Vascular Surgery Vascular Quality Initiative.
Carotid endarterectomy (CEA) is among the most commonly performed vascular procedures. Some have suggested worse outcomes with contralateral internal carotid artery (ICA) occlusion. We compared patients with and patients without contralateral ICA occlusion using the Society for Vascular Surgery Vascular Quality Initiative database. Deidentified data were obtained from the Vascular Quality Initiative. Patients with prior ipsilateral or contralateral CEA, carotid stenting, combined CEA and coronary artery bypass graft, or <1-year follow-up were excluded, yielding 1737 patients with and 45,179 patients without contralateral ICA occlusion. Groups were compared with univariate tests, and differences identified in univariate testing were entered into multivariate models to identify independent predictors of outcomes and in particular whether contralateral ICA occlusion is an independent predictor of outcomes. Patients with contralateral ICA occlusion were younger and more likely to be smokers; they were more likely to have chronic obstructive pulmonary disease, preoperative neurologic symptoms (56% vs 47%), nonelective CEA (16% vs 13%), and shunt placement (75% vs 53%; all P < .001). The 30-day ipsilateral stroke risk was 1.3% with vs 0.7% without contralateral ICA occlusion (P = .004). The 30-day and 1-year survival estimates were 99.0% ± 0.5% and 94.1% ± 1.1% with vs 99.6% ± 0.1% and 96.0% ± 0.2% without contralateral ICA occlusion (log-rank, P < .001). Logistic regression analysis identified prior neurologic event (P = .046), nonelective surgery (P = .047), absence of coronary artery disease (P = .035), and preoperative angiotensin-converting enzyme inhibitor use (P = .029) to be associated with 30-day ipsilateral stroke risk, but contralateral ICA occlusion remained an independent predictor in that model (odds ratio, 2.29; P = .026). However, after adjustment for other factors (Cox proportional hazards), risk of ipsilateral stroke (including perioperative) during follow-up was not significantly greater with contralateral ICA occlusion (hazard ratio, 1.21; P = .32). Results comparing propensity score-matched cohorts mirrored those from the larger data set. This study demonstrates likely clinically insignificant differences in early stroke or death in comparing CEA patients with and those without contralateral ICA occlusion. After adjustment for other factors, contralateral ICA occlusion was not associated with a greater risk of ipsilateral stroke (including perioperative) in longer follow-up. Mortality was greater with contralateral ICA occlusion, and this difference was more pronounced at 1 year despite younger age of the contralateral ICA occlusion group. CEA risk remains low even in the presence of contralateral ICA occlusion and appears to be explained at least in part by other factors. CEA should still be considered appropriate in the face of contralateral ICA occlusion.


Structure and expression of the hepatitis B virus genome.
By fusion of the hepatitis B virus (HBV) surface antigen (HBsAg) gene to that of the E. coli lac Z gene carried by a phage lambda derivative, expression of HBsAg antigenic determinants was obtained and carried by a 138,000 dalton fusion polypeptide. Such a protein could be ultimately useful for second generation vaccine production. HBsAg gene expression was studied in eukaryotic cells using the mouse L cell (tk-) system. Cotransformation using a plasmid carrying two copies of the HBV genome in a tandem, head-to-tail arrangement (pCP10) and the cloned HSV-1 tk gene resulted in the excretion of 22 nm HBsAg particles in the supernatant. No other HBV markers were detected. These particles possess the same characteristics as the human serum particles (morphology, diameter, density, antigenicity). The purified HBsAg particles from L cells were found to be highly immunogenic in mice. HBV mRNA transcripts from these cells were analysed by Northern blotting. A major species of 2,300 bases was detected. This was mapped on the genome by hybridization with subgenomic fragments and in the L cell system using a series of plasmid derivatives carrying insertions at specific sites in the HBV genome and assaying for HBsAg expression. Thus the HBsAg gene promotor was localized between positions 2,400-2,800. Indeed there is only one TATA like sequence in this region, starting at position 2,776.


Capsaicin inhibits A7r5 cell senescence via the mitochondrial carrier protein Slc25a12.
Aging of vascular smooth muscle cells (VSMCs) is the principal factor responsible for the loss of vascular function, and continuous exposure to high glucose is one of the key factors contributing to the aging of VSMCs. This study established a high glucose-induced senescence model of the A7r5 cell line and used transcriptome sequencing to screen the regulatory target genes of high glucose-induced cellular senescence. The study revealed that the expression of the Slc25a12 gene, which belongs to the solute carrier family 25 member 12, was notably reduced following damage caused by high glucose levels. This inhibition was shown to cause mitochondrial malfunction and cellular senescence. The encoded product of the Slc25a12 gene is a mitochondrial carrier protein that binds to calcium and aids in transporting aspartate for glutamate exchange within the inner mitochondrial membrane. Mitochondrial dysfunction compromises the cell's capacity to resist oxidation and repair damage, and is an inherent element in hastening cellular aging. Moreover, our findings validated that the transient receptor potential vanilloid 1 (TRPV1) agonist capsaicin hindered the decrease in Slc25a12 expression, prevented mitochondrial dysfunction, and blocked cellular senescence. Could the regulation of Slc25a12 expression by capsaicin restore cellular mitochondrial function and restrict senescence? In vitro tests have verified that interference with A7r5 Slc25a12 noticeably diminishes capsaicin's effectiveness in repairing mitochondrial function and inhibiting senescence. The findings indicate that capsaicin delays mitochondrial dysfunction and therefore hinders cellular senescence by regulating the mitochondrial membrane protein Slc25a12 in the A7r5 cell line.


Deciphering the origins of molecular toxicity of combretastatin A4 and its glycoconjugates: interactions with major drug transporters and their safety profiles in vitro and in vivo.
Cellular uptake and transport mechanisms directly correlate with the drug-like profiles of lead compounds. To decipher the molecular origin of the toxicity of combretastatin A4 (CA4), an important microtubule targeting agent, we investigated the interactions between CA4 and six key drug transporters, namely hOAT1, hOAT3, hOCT1, hOCT2, hOATP1B3, and hOATP2B1. Three combretastatin-based glycoconjugates, namely Glu-CA4, Man-CA4, and Gal-CA4 with glucose, mannose, and galactose respectively, were synthesized and their in vitro and in vivo biological characteristics were evaluated. CA4 exhibited significant inhibition against hOAT3 and hOATP2B1, moderate inhibition of hOAT1 and hOCT2, and weak inhibitory effects on hOCT1 and hOATP1B3. Compared to CA4, the inhibitory activities of Glu-CA4 on the six transporters were minimal. The glycoconjugates were found to have a superior safety profile with their maximum tolerated dose (MTD) values exhibiting a 16-34-fold increase compared to CA4. Given the drawbacks of CA4, the enhanced solubility and safety profiles of CA4 glycoconjugates augur well for further investigation into these intriguing candidates' in vivo efficacy.


Vitamin D measured in maternal serum and offspring neurodevelopmental outcomes: a prospective study with long-term follow-up.
Vitamin D is obtained from dietary sources and synthesized in the skin during exposure to ultraviolet B radiation in sunlight. During pregnancy, vitamin D is transported from mother to fetus through the placenta in the form of 25-hydroxyvitamin D [25(OH)D]. There is evidence that vitamin D influences neuronal differentiation, endocrine functions, and fetal brain growth. Animal studies indicate alterations in the offspring brain as a consequence of vitamin D deficiency during pregnancy. In humans, maternal vitamin D insufficiency has been linked to impaired child language development. Using data from a prebirth cohort with up to 22 years of follow-up, we examined the association of vitamin D status with proxies of offspring neurodevelopmental outcomes. During 1988-1989, pregnant women were recruited for the DaFO88 cohort (n = 965) in Aarhus, Denmark. Maternal concentrations of 25(OH)D were quantified in serum from week 30 of gestation via the LC-MS/MS method (n = 850). Offspring were followed up through national registries until the age of 22 years. We evaluated the association of the maternal concentration of 25(OH)D with offspring neurodevelopmental outcomes defined as first admission diagnosis or prescription of medication for (1) ADHD, (2) depression, and (3) scholastic achievement based on the mean grade on standardized written examinations in the 9th grade (final exams after 10 years of compulsory school in Denmark). Maternal concentrations of 25(OH)D were higher compared to current levels (median 76 nmol/l; 5th to 95th percentiles 23-152). There was a direct association between maternal vitamin D status and offspring depression (p(trend) = 0.01); for ADHD there was no association. Scholastic achievement was slightly higher for offspring of mothers with a vitamin D status in the range of >50-125 nmol/l, but this nonlinear association was not statistically significant. Our analyses based on biomarker measurement of 25(OH)D from a cohort of 850 pregnant women combined with long-term follow-up showed no support for a beneficial fetal programming effect of vitamin D status with regard to behavioral and affective disorders and scholastic achievement.


A member of the peptidase M48 superfamily of Porphyromonas gingivalis is associated with virulence in vitro and in vivo.
In vivo-induced antigen technology was previously used to identify 115 genes induced in Porphyromonas gingivalis W83 during human infection. One of these, PG2197, a conserved hypothetical protein which has homology to a Zn-dependent protease, was examined with respect to a role in disease. The expression of PG2197 in human periodontitis patients was investigated, but as there is increasing evidence of a direct relationship between P. gingivalis and cardiovascular disease, a mutation was constructed in this gene to also determine its role in adherence, invasion, and persistence within human coronary artery endothelial cells (HCAEC) and neutrophil killing susceptibility. Plaque samples from 20 periodontitis patients were analyzed by real-time PCR, revealing that PG2197 was expressed in 60.0% of diseased sites compared to 15.8% of healthy sites, even though P. gingivalis was detected in equal numbers from both sites. The expression of this gene was also found to be up-regulated in microarrays at 5 and 30 min of invasion of HCAEC. Interestingly, a PG2197 mutant displayed increased adherence, invasion, and persistence within HCAEC when compared to the wild-type strain. This gene appears to be important for the virulence of P. gingivalis, both in vivo and in vitro.


Predicting falls within the elderly community: comparison of postural sway, reaction time, the Berg balance scale and the Activities-specific Balance Confidence (ABC) scale for comparing fallers and non-fallers.
Simple reaction time, the Berg balance scale, the Activities-specific Balance Confidence (ABC) scale and postural sway were studied in order to determine cut-off scores as well as develop a model used in the prevention of fallers within the elderly community. One hundred and twenty-five subjects, 45 fallers and 80 non-fallers were evaluated throughout the study and results indicated that non-fallers have significantly faster reaction times, have higher scores on the Berg balance scale and the ABC scale as well as sway at slower frequencies when compared to fallers. Furthermore, all risk factors were subsequently entered into a logistic regression analysis and results showed that reaction time, the total Berg score and the total ABC score contributed significantly to the prediction of falls with 89% sensitivity and 96% specificity. A second logistic regression was carried out with the same previous variables as well as all questions of the Berg and ABC scales. Results from the logistic analysis revealed that three variables were associated with fall status with 91% sensitivity and 97% specificity. Results from the following study would seem rather valuable as an assessment tool for health care professionals in the identification and monitoring of potential fallers within nursing homes and throughout the community.


Effect of Lactic Acid Bacteria Strains on the Growth and Aflatoxin Production Potential of Aspergillus parasiticus, and Their Ability to Bind Aflatoxin B1, Ochratoxin A, and Zearalenone in vitro.
The increased consumption of plant-based foods has intensified the concern related to mycotoxin intoxication. This study aimed to investigate the effect of selected lactic acid bacteria (LAB) strains on the growth of Aspergillus parasiticus NRRL 2999 and its production of aflatoxin (AF). The ability of the heat-killed (100°C for 1 h) LAB strains to bind aflatoxin M1 (AFM1) in milk and aflatoxin B1 (AFB1), ochratoxin A (OTA), and zearalenone (ZEN) in potassium phosphate buffer (PPB) was also evaluated in vitro. Ten LAB strains were tested individually, by inoculating them simultaneously with the fungus or after incubation of the fungus for 24 or 48 h at 25°C. Double layer yeast extract sucrose (YES) agar, de Man Rogosa and Sharpe (MRS) agar, and YES broth were incubated for 7 days at 25°C to follow the development of the fungus. Levilactobacillus spp. 3QB398 and Levilactobacillus brevis 2QB422 strains were able to delay the growth of A. parasiticus in YES broth, even when these strains were inoculated 24 h after the fungus. The inhibitory effect of these LAB strains was confirmed by the reduction of fungus colony size, suggesting dominance of LAB by competition (a Lotka-Voltera effect). The production of AFB1 by A. parasiticus was inhibited when the fungus was inoculated simultaneously with Lactiplantibacillus plantarum 3QB361 or L. plantarum 3QB350. No AFB1 was found when Levilactobacillus spp. 2QB383 was present, even when the LAB was inoculated 48 h after the fungus. In binding studies, seven inactivated LAB strains were able to promote a reduction of at least 50% the level of AFB1, OTA, and ZEN. This reduction varied depending on the pH of the PPB. In milk, however, only two inactivated LAB strains were able to reduce AFM1, with a reduction of 33 and 45% for Levilactobacillus spp. 3QB398 (Levilactobacillus spp.) and L. brevis 2QB422, respectively. Nevertheless, these results clearly indicate the potential of using LAB for mycotoxin reduction.


Stimulation of chlororespiration by heat and high light intensity in oat plants.
High irradiance and moderate heat inhibit the activity of the photosynthetic apparatus of oat (Avena sativa L.) leaves. The incubation of oat leaves under high light intensity in conjunction with high temperatures strongly decreased the maximal quantum yield of photosystem (PS) II, indicating the close synergistic effect of both stress factors on PS II inhibition and the subsequent irreversible damage to the photosynthetic apparatus. The PS I A/B protein levels remained similar to control values in leaves incubated under high light intensity or moderate heat, and decreased only when both stress factors were simultaneously applied. Immunoblot analysis of thylakoid membranes using specific antibodies raised against the NDH-K subunit of the thylakoidal NADH dehydrogenase complex (NADH DH) and against plastid terminal oxidase (PTOX) revealed an increase in the amount of both proteins in response to high light intensity and/or heat treatments. In addition, these stress treatments were seen to stimulate the activity of electron donation by NADPH and ferredoxin to plastoquinone, the PTOX activity in plastoquinone oxidation and the NADH DH activity in thylakoid membranes. Incubation with n-propyl gallate (an inhibitor of PTOX) inhibited the increase of NDH-K and PTOX levels under high light intensity and heat, and slightly stimulated the activity of electron donation by NADPH and ferredoxin to plastoquinone. Antimycin A (an inhibitor of cyclic electron flow) increased the NADH DH activity and preserved the levels of NDH-K and PTOX in thylakoid membranes from leaves incubated under high light intensity and heat. The up-regulation of the PTOX and the thylakoidal NADH DH complex under these stress conditions supports a role for chlororespiration in the protection against high irradiance and moderate heat.


Effects of crowding on the thermal stability of heterogeneous protein solutions.
Crowding can substantially affect the transition of a protein between its native (N) and unfolded (U) states via volume exclusion effects. Also, it influences considerably the aggregation (A) of unfolded proteins. To examine the details, we developed an approach for computing the kinetic rates of the process N <--> U --> A in which the concentration of the protein is explicitly taken into account. We then compute the relative change with temperature of the protein denaturation for various fractional volume occupancies and partition of proteins in solution. The analysis indicates that, in protein solutions in which the average distance between proteins is comparable with the radius of gyration of an unfolded protein, steric effects increase the stability of the proteins which are in compact, native states. In heterogeneous protein solutions containing various types of proteins with different thermal stabilities, the unfolding of the most thermolabile proteins will increase the stability of the other proteins. The results shed light on the way proteins change the thermal stability of a cell as they unfold and aggregate. This study may be valuable in questions related to the dynamics of thermal injuries.


Role of PknB kinase in antibiotic resistance and virulence in community-acquired methicillin-resistant Staphylococcus aureus strain USA300.
The regulation of cellular processes by eukaryote-like serine/threonine kinases is widespread in bacteria. In the last 2 years, several studies have examined the role of serine/threonine kinases in Staphylococcus aureus on cell wall metabolism, autolysis, and virulence, mostly in S. aureus laboratory isolates in the 8325-4 lineage. In this study, we showed that the pknB gene (also called stk1) of methicillin-resistant S. aureus (MRSA) strain COL and the community-acquired MRSA (CA-MRSA) strain USA300 is involved in cell wall metabolism, with the pknB mutant exhibiting enhanced sensitivity to beta-lactam antibiotics but not to other classes of antibiotics, including aminoglycosides, ciprofloxacin, bactrim, and other types of cell wall-active agents (e.g., vancomycin and bacitracin). Additionally, the pknB mutant of USA300 was found to be more resistant to Triton X-100-induced autolysis and also to lysis by lysostaphin. We also showed that pknB is a positive regulator of sigB activity, resulting in compromise in its response to heat and oxidative stresses. In association with reduced sigB activity, the expression levels of RNAII and RNAIII of agr and the downstream effector hla are upregulated while spa expression is downmodulated in the pknB mutant compared to the level in the parent. Consistent with an enhanced agr response in vitro, virulence studies of the pknB mutant of USA300 in a murine cutaneous model of infection showed that the mutant was more virulent than the parental strain. Collectively, our results have linked the pknB gene in CA-MRSA to antibiotic resistance, sigB activity, and virulence and have highlighted important differences in pknB phenotypes (virulence and sigB activity) between laboratory isolates and the prototypic CA-MRSA strain USA300.


Cost of Oncology Drugs in the Middle-Eastern Country of Lebanon: An Update (2014-2016).
This study aims to evaluate trends in the increasing costs of oncology drugs procured by the Lebanese Ministry of Public Health (MOPH) between 2014 and 2016 and to assess the impact of the introduction in mid-2015 of new immunotherapy drugs for the treatment of lung cancer on the overall and specific costs of that treatment. A secondary analysis of data from the MOPH Cancer Drug Scientific Committee data base was conducted using a total of 18,133 cancer files between 2014 and 2016. Over the 3-year period, about $140 million (USD) was spent on cancer drugs by the MOPH free cancer drug dispensing program. The expenditures increased by 27% after immunotherapy was phased in. The average cost of drugs per patient per year measured across all cancer types increased from $7,000 in 2014 to $8,400 in 2016. Trastuzumab, approved for treating human epidermal growth factor receptor 2-positive breast cancer ranked first in total expenditures for 2014-2015. By 2016, two new immunotherapy drugs had topped the list: pembrolizumab ranked first and nivolumab ranked third, representing 64% of the total cost of lung cancer treatment and approximately 19% of the total yearly budget; beneficiaries represented only 3% of all patients. This update documents the increasing financial impact of newer cancer drugs on the procurement process in the middle-income country of Lebanon. The trend is aligned with the financial burden of cancer drugs worldwide, which calls for a collaborative global response to this crisis.


Pancreatic glucokinase is activated by insulin-like growth factor-I.
Glucokinase (GK) plays a key role in the regulation of glucose use and glucose-stimulated insulin secretion in pancreatic islet cells. Gene targeting of the IGF-I receptor down-regulated pancreatic islet GK activity. That finding prompted us to examine the potential mechanism that may control GK gene activity using an islet cell line, INS-1, known to express IGF-I receptor. Exposure of these cells to IGF-I induced GK protein expression and activity of the enzyme in a dose-dependent manner. In addition, IGF-I induced activity of a reporter construct containing the GK promoter in parallel with the effect on endogenous GK mRNA levels. The stimulatory effect of IGF-I on GK promoter activity was abrogated by wortmannin and LY294002, specific inhibitors of phosphatidylinositol 3-kinase. Exposure of cells to IGF-I elicited a rapid phosphorylation of Akt and FoxO1, a known target of Akt signaling. Constitutively active Akt stimulates the activity of the GK promoter, and a dominant-negative mutant of Akt or mutagenesis of a FoxO1 response element in the GK promoter abolished the ability of IGF-I to stimulate the promoter activity. Furthermore, cell knockdown of FoxO1 with small interfering RNA disrupted the effect of IGF-I on GK expression. These results demonstrate that the phosphatidylinositol 3-kinase/Akt/FoxO1 pathway contributes to the regulation of GK gene expression in response to IGF-I stimulation.


Peripheral blood gene expression analysis in intestinal transplantation: a feasibility study for detecting novel candidate biomarkers of graft rejection.
We investigated the putative candidate biomarkers of graft rejection in peripheral blood of intestinal transplant patients. Peripheral blood gene expression analysis was performed in intestinal transplant patients. The results were matched with concurrent graft biopsies using bioinformatics. Peripheral blood samples (n=11), of 3 adult patients [transplant day (n=1), no rejection (n=1), minimal rejection (n=2), mild rejection (n=5) and severe rejection (n=2)] were collected. Bioinformatics: Enrichment Analysis: The three most affected pathways differentially expressed in rejection versus a pool of healthy volunteers were related to protein translation: translation initiation, translation elongation termination, and translation in mitochondria, with p-values for all rejection stages in all patients in the 10-4 to 10-18 range. No significant enrichment was observed for these categories in the day of transplant sample. In addition to translation, significant enrichment of several immune response categories was observed in rejection samples. Subsequent gene set enrichment analysis verified these results. The level of enrichment was very high (p-values of 10-5-10-60) and increased with the level of rejection in all patients. Genes significantly down-regulated in translation related gene sets included ribosomal proteins RPL13A, RP L22, RPS23, RPL13 and RPL10A, that could be used as potential biomarkers for future experiments. In this pilot study we found a list of genes (involved in translation) significantly downregulated in the peripheral blood of three intestinal transplant patients during rejection. These results will be verified in further studies with increased number of patients and with isolation of peripheral blood subpopulations.


[Cutaneous innervation: anatomy and physiology. Neuropeptides].
The skin is involved in the tactile sensations and hydric and thermic regulation. This structure contains immunological cells with inflammatory activity in addition to several bioactive molecules. This review will focus primarily on the relation of cutaneous innervation in these processes and its capacity to release neuropeptides as functional molecules. This paper will discuss the organization of the motor and sensitive fibres, the biosynthesis, molecular modifications, regulation and the functions of the best well known neuropeptides. The specific function of these molecules in the neurogenic inflammation and how the nervous system can influence in several skin diseases is explained. In conclusion, the cutaneous innervation can affect processes such as inflammation, cellular proliferation and healing. This possibility is likely to be mediated by neuropeptides. In the future, a growing list of neuropeptides will be compiled and the precise role in cutaneous physiology will be known, but the most important of this knowledge is probably the repercussion in the therapies of some cutaneous diseases.


Comparison of Salmonella serovar isolation and antimicrobial resistance patterns from porcine samples between 2003 and 2008.
Food-borne Salmonella infections can produce symptoms from mild gastroenteritis to severe systemic disease and death, representing an important public health issue in U.S. livestock and livestock products, which have been implicated as frequent sources of Salmonella contamination. Concerns have been raised about the spread of antibiotic resistance in Salmonella strains, particularly those that originate from food animal sources, as a result of prophylactic and therapeutic antimicrobial use in these species. Longitudinal comparisons of Salmonella serovars isolated from porcine tissues submitted to the Iowa State University Veterinary Diagnostic Laboratory in 2003 and 2008 were conducted to evaluate changes in serovar dynamics and antimicrobial resistance. Incidence of recovered group C Salmonella enterica serovar Choleraesuis var. Kunzendorf decreased between 2003 and 2008, while recovery of group B strains Salmonella Typhimurium var. 5-(formerly, Copenhagen), Salmonella Agona, Salmonella Derby, Salmonella Heidelberg, and Salmonella Typhimurium increased. Significant changes in resistance interpretation were seen in Salmonella Derby with regard to spectinomycin and sulfadimethoxine; in Salmonella Heidelberg with regard to florfenicol, spectinomycin, and sulfadimethoxine; and in Salmonella Choleraesuis var. Kunzendorf, Salmonella Typhimurium, Salmonella Typhimurium var. 5-, and Salmonella Agona with regard to spectinomycin. Only 2 of 293 isolates in 2003 and 5 of 395 isolates in 2008 were resistant to enrofloxacin. Utilizing antibiotics approved for use in food animals to evaluate antimicrobial resistance provides more specific information on the selection pressure exerted on Salmonella populations through the use of these drugs.


The association of hospital characteristics and quality improvement activities in inpatient medical services.
Quality of U.S. health care has been the focus of increasing attention, with deficiencies in patient care well recognized and documented. However, relatively little is known about the extent to which hospitals engage in quality improvement activities (QIAs) or factors influencing extent of QIAs. To identify 1) the extent of QIAs in Veterans Administration (VA) inpatient medical services; and 2) factors associated with widespread adoption of QIAs, in particular use of hospitalists, non-physician providers, and extent of goal alignment between the inpatient service and senior managers on commitment to quality. Cross-sectional, descriptive study of QIAs using a survey administered to Chiefs of Medicine (COM) at all 124 VA acute care hospitals. We conducted hierarchical regression, regressing QIA use on facility contextual variables, followed by use of hospitalists, non-physician providers, and goal alignment/quality commitment. Outcome measures pertained to use of a set of 27 QIAs and to three dimensions--infrastructure, prevention, and information gathering--that were identified by factor analysis among the 27 QIAs overall. Survey response rate was 90 % (111/124). Goal alignment/quality commitment was associated with more widespread use of all four QIA categories [infrastructure (b = 0.42; p < 0.001); prevention (b = 0.24; p < 0.001); information gathering (b = 0.28; p = <0.001); and overall QIA (b = 0.31; p < 0.001)], as was greater use of hospitalists [infrastructure (b = 0.55; p = 0.03); prevention (b = 0.61; p < 0.001); information gathering (b = 0.75; p = 0.01); and overall QIAs (b = 0.61; p < 0.001)]; higher occupancy rate was associated with greater infrastructure QIAs (b = 1.05, p = 0.02). Non-physician provider use, hospital size, university affiliation, and geographic region were not associated with QIAs. As hospitals respond to changes in healthcare (e.g., pay for performance, accountable care organizations), this study suggests that practices such as use of hospitalists and leadership focus on goal alignment/quality commitment may lead to greater implementation of QIAs.


New U.S. guidelines for prescribing dental radiographs--a critical review.
Most national dental organizations have approved recommendations which set out principles to guide the profession in its use of diagnostic radiation. In 1987, the Center for Device and Radiological Health (an agency of the United States government) published detailed guidelines. This significant initiative should encourage and accelerate the practical application of these principles. In this article, we identify and discuss what we determine to be a major flaw in the guidelines regarding radiography for children in the transitional dentition stage. As well, we have suggested revisions to the guidelines to eliminate some less serious flaws which we believe add unnecessary complexity to the chart.


Selection and characterization of ColE1 plasmid mutants that exhibit altered stability and replication.
This report describes a method for isolating mutants of plasmid ColE1 that exhibit unstable maintenance and altered replication characteristics. It also describes the initial characterization of four mutants isolated by that method. A chimeric plasmid, pHSG124, containing a ColE1 derivative and a temperature-sensitive replication derivative of pSC101 was mutagenized in vitro, using hydroxylamine. By adjusting the growth conditions of transformants containing the mutagenized chimeric deoxyribonucleic acid, it was possible to rapidly screen colonies and identify those that had a high probability of carrying ColE1 mutants that exhibit unstable maintenance. Of those mutants, some exhibited altered copy number or accumulated catenated structures. Evidence is presented which suggests that the mutations in three of the mutants are probably located in the HaeII A fragment of ColE1.


Reconstitution of wild-type or mutant telomerase activity in telomerase-negative immortal human cells.
Telomere shortening in human somatic cells and telomere maintenance in most human immortal cell lines and tumours correlate respectively with the absence and presence of telomerase, the enzyme that synthesizes telomeric DNA de novo . However, approximately 30% of in vitro immortalized human cell lines do not express this enzyme and maintain telomeres by an alternative pathway (ALT) that may also operate in some tumours. Human telomerase is a reverse transcriptase comprising minimally an RNA subunit (hTER) and a catalytic protein moiety (hTERT). Normal somatic cells retain expression of hTER but not of hTERT, and can be converted to a telomerase-positive phenotype by ectopic expression of the catalytic protein. We similarly have restored enzymatic activity to those ALT cell lines that retain hTER expression. We also report that in those ALT cells that are hTER negative, reintroduction of both hTER and hTERT is necessary and sufficient for conversion to telomerase positivity. Moreover, transfection of these cells with hTERT in conjunction with hTERs with a mutated template results in the expression of an enzyme with altered specificity. Reconstitution of telomerase activity in ALT cells, particularly an activity capable of synthesizing mutant telomeric DNA, may be exploited for the study of the ALT mechanism and its interaction with the telomerase-dependent pathway, and for assessing the effects of mutant telomeres on cell viability.


Goblet cell carcinoids: characteristics of a Danish cohort of 83 patients.
Appendiceal goblet cell carcinoids (GCCs) exhibit neuroendocrine and adenocarcinoma features. Analysis of demography, pathology, prognostic markers, treatment and survival in 83 GCC patients (f/m: 56/27) diagnosed 1992-2013. Median age for f/m was 59/58 years, respectively, and similar for localized and disseminated disease. At diagnosis 54 patients had localized appendiceal disease (f/m: 29/25). According to TNM 24% had Stage I, 70% had Stage II and 6% had Stage III. Twenty-nine patients had disseminated disease (f/m: 27/2). Chromogranin A, synaptophysin and p53 were positive in >90%. Serotonin was positive in 70%. Median Ki67 index was 32% (6-75%) and higher in Tang group C (50%) compared to group A (30%; p<0.0001), and group B (30%; p<0.004). All patients had surgery. Sixty-three (76%) had radical resections including all patients with localized disease. Median OS was 83 months. The 1-, 5- and 10-year survival rates were 90%, 58%, and 38%, respectively. For localized disease OS was 164 months and 1-, 5- and 10-year survival rates were 100%, 80%, and 55%, respectively. For disseminated disease OS was 19 months and 1-, 5- and 10-year survival rates were 73%, 18% and 6%, respectively. The 1-, 5- and 10 year-survival rates for f/m were 87%/96%, 49%/76% and 31%/57%, respectively (p = 0.02). According to the Tang classification group A, B, and C OS was 118, 83 and 20 months, respectively (p = 0.0002). The Tang classification was found to be a significant prognostic factor, while the Ki67 index was not. Localized GCCs occurred equally in males and females, while disseminated GCCs were mostly seen in females. Median age of patients with localized disease and disseminated disease was identical. Cox regression analysis found Stage IV, focally positive synaptophysin and non-radical surgery as strongest negative prognostic factors.


Working With Infertile Couples Seeking Assisted Reproduction: An Interpretative Phenomenological Study With Infertility Care Providers.
Although most studies investigated the impact of infertility and its treatment on the couple, a small body of evidence suggested that infertility care providers may experience different sources of stress related for instance to excessive workload, the complexity of the technique, and relational difficulties with patients. The current study aimed at providing further insight into the understanding of the subjective experience of infertility care providers by highlighting their feelings and emotions, personal meanings, challenges, and opportunities. Following the methodological guidelines of Interpretative Phenomenological Analysis, we conducted individual semi-structured interviews with 23 members of two different fertility units. Interviews were audiotaped and transcribed verbatim. Textual analysis was then conducted to identify emerging dominant themes and subthemes. Three main themes were extracted: (i) dealing with infertile patients and their specificities, (ii) performing assisted reproductive technology (ART), (iii) being part of a team. These themes related to participants experiencing: (i) difficulties in establishing an empathic connection and communicating with couples, such that women were sometimes perceived as "particular patients" and men as poorly involved in the process; (ii) difficulties in dealing with a complex procedure in which errors are not allowed (as reported by embryologists), with a growing number of women aged > 40 seeking assisted reproduction, despite the risks for their health; (iii) being part of a team as a resource, although the huge amount of time spent together can involve conflicts and organizational problems. These findings suggested that patients' overpersistence (rather than just dropout) represents an important source of stress for infertility care providers. At the same time, the concept of particular or difficult patient derives from the combination of multiple factors, including providers' own history and subjective experience. The presence of mental health professionals in fertility units is essential to help providers improve the quality of doctor-patient communication and relieve the stress related to organizational issues and conflicts.


Polyprotein processing and intermolecular interactions within the viral replication complex spatially and temporally control norovirus protease activity.
Norovirus infections are a major cause of acute viral gastroenteritis and a significant burden on global human health. A vital process for norovirus replication is the processing of the nonstructural polyprotein by a viral protease into the viral components required to form the viral replication complex. This cleavage occurs at different rates, resulting in the accumulation of stable precursor forms. Here, we characterized how precursor forms of the norovirus protease accumulate during infection. Using stable forms of the protease precursors, we demonstrated that all of them are proteolytically active in vitro, but that when expressed in cells, their activities are determined by both substrate and protease localization. Although all precursors could cleave a replication complex-associated substrate, only a subset of precursors lacking the NS4 protein were capable of efficiently cleaving a cytoplasmic substrate. By mapping the full range of protein-protein interactions among murine and human norovirus proteins with the LUMIER assay, we uncovered conserved interactions between replication complex members that modify the localization of a protease precursor subset. Finally, we demonstrate that fusion to the membrane-bound replication complex components permits efficient cleavage of a fused substrate when active polyprotein-derived protease is provided in trans These findings offer a model for how norovirus can regulate the timing of substrate cleavage throughout the replication cycle. Because the norovirus protease represents a key target in antiviral therapies, an improved understanding of its function and regulation, as well as identification of interactions among the other nonstructural proteins, offers new avenues for antiviral drug design.


Gold nanoparticles - an optical biosensor for RNA quantification for cancer and neurologic disorders diagnosis.
The objective of this study is to develop a facile tool for the absolute detection and quantification of nucleic acid transcripts, using a gold nanoparticle-based optical biosensor. Topoisomerase 1 (TOP1) and tyrosyl DNA phosphodiesterase 2 (TDP2) were among the nucleic acid transcripts of choice due to their role as genomic instability biomarkers and their implication in various cancers and neurologic disorders. This opens the door to develop a simple tool that can be used for diagnosing and monitoring treatment response for such diseases, overcoming the requirements for high cost, time, and complexity of the existing technologies for the absolute quantification of transcripts of interest. The TOP1 and TDP2 mRNA transcripts were first captured specifically using magnetic nanoparticles that were functionalized with TOP1- and TDP2-specific probes, respectively. The captured mRNA was then directly detected and quantified using the gold aggregating gold (GAG) assay, without the need for amplification as in existing technologies used for the quantification of transcripts. A linear correlation exists between the GAG assay and the qPCR for the quantification of the TOP1 and TDP2 mRNA transcripts (101-104 copies). The detection limit of the GAG assay in mRNA quantification was up to 10 copies per reaction. Wild-type and TDP2-deficient cell lines confirmed the assay specificity and reproducibility in distinguishing between different transcripts. The GAG assay can be utilized as an inexpensive, rapid, simple, and sensitive tool for the absolute quantification of RNA for different applications, instead of the laborious, expensive, and sophisticated real-time PCR.


A putative heat-labile enterotoxin expressed by strains of Aeromonas media.
Fifteen isolates of Aeromonas media (seven from diarrhoeal stools, four from water and four from superficial skin ulcers of catfish) were examined for enterotoxin production. Ten of these isolates (six diarrhoeal, one from water and three from fish) caused accumulation of fluid in the initial rabbit ileal loop (RIL) tests. Isolates from diarrhoeal stools and fish caused relatively more fluid accumulation than those from water. Those strains that caused little or no fluid accumulation in the initial experiments became enterotoxin producers after one passage through RILs, regardless of source, and also showed gradual enhancement of fluid outpouring after each subsequent passage. Inocula of c. 1 x 10(4) viable cells and 0.25 ml of culture filtrate (CF) caused fluid accumulation similar to that of toxigenic Vibrio cholerae 569B. The enterotoxic factor(s) were inactivated when held at 56 degrees C for 20 min or 65 degrees C for 10 min and showed biological activity over a wide range of pH values. These results suggest that strains of A. media, whether from diarrhoeal stools, water or infected fish, are potentially enterotoxigenic and may have the potential to produce a heat-labile and pH-stable diarrhoeagenic factor in the same way as other known heat-labile and pH-stable enterotoxins.


Predictions without templates: new folds, secondary structure, and contacts in CASP5.
We present the assessment of CASP5 predictions in the new fold category. For coordinate predictions, we considered five targets with new folds and eight lying on the fold recognition borderline. We performed detailed visual and numerical comparisons between predicted and experimental structures to assess prediction accuracy. The two procedures largely agreed, but the visual inspection identified instances where metrics, such as GDT_TS, ranked what we considered incorrect predictions highly. We found the quality of the best predictions to be very good: for nearly every target at least one group predicted a structure close to the correct one. However, selection of the best of five models is still problematic. The group of David Baker once again proved to be best overall, with many individual highlights. However, high quality and consistency were also seen from others, suggesting that the community is moving toward general procedures to predict accurate structures for proteins showing no resemblance to anything seen before. Predictions for secondary structure showed at best limited progress since CASP4. The number of targets is probably too small to spot differences in performance between methods, suggesting that such predictions might be better evaluated with schemes involving more proteins. For contact predictions, accuracies are still low, although there were several instances of accurate and useful contacts predicted de novo, and new approaches hint at future progress.


Correlation between presence of primary iris- and cilliary body cysts and intraocular pressure.
The objective of this study was to examine the correlation between the presence of primary iris-ciliary cysts and the intraocular pressure. Sixty patients with short-sightedness undergoing routine examination for laser vision correction in our hospital in 2003 were enrolled. Patients with known high intraocular pressure and risk of glaucoma were excluded from the study. A total of 119 eyes were examined by the Ultrasound Biomicroscope (UBM), and the presence of the primary iris-ciliary cysts was confirmed. Intraocular pressure was measured by using a blowing tonometer for each eye in triplicate. Through Pentacam correction of intraocular pressure using the Ehlers formula, the influence of the thickness of central cornea on intraocular pressure was excluded. Among all participants, 62 eyes (52.1%) were with high myopia, 57 eyes (47.9%) with low and moderate myopia, 27 eyes (22.7%) with single cyst, 20 eyes (16.8%) with multiple cysts, and 72 eyes (60.5%) were free from cysts. Moreover, the intraocular pressure was found within the normal range in 72 eyes (60.5%), and abnormally high in 47 eyes (39.5%). Our results showed that the presence of primary iris-ciliary cysts and the intraocular pressure were positively correlated, with a correlation coefficient of 0.235 (p = 0.01). These findings may prove useful for prediction and screening of high intraocular pressure.


Drug-loaded carbon nanohorns: adsorption and release of dexamethasone in vitro.
Single-wall carbon nanohorns (SWNHs) are recently discovered nanostructured spherical aggregates of graphitic tubes. The unique physicochemical properties of SWNHs, including their large surface area, suggest their possible utility as carriers in drug delivery systems. Here we investigated the in vitro binding and release of the antiinflammatory glucocorticoid dexamethasone (DEX) by as-grown SWNHs and their oxidized form, oxSWNHs. Adsorption analyses using [3H]-DEX determined the amount of DEX adsorbed by oxSWNHs to be 200 mg for each gram of oxSWNHs in 0.5 mg/mL of DEX solution, which was approximately 6 times larger than that obtained for as-grown SWNHs. Adsorption kinetics indicated that oxSWNHs had higher affinity for DEX than as-grown SWNHs. Treatment of oxSWNHs at 1200 degrees C under H2, which removed the oxygen-containing functional groups on oxSWNHs, did not diminish the high affinity for DEX, suggesting that oxygen-containing functional groups have little contribution for the affinity. DEX-oxSWNH complexes exhibited sustained release of DEX into phosphate-buffered saline (pH 7.4) at 37 degrees C and more rapid biphasic release into culture medium. The biological integrity of the released DEX form was confirmed by activation of glucocorticoid response element-driven transcription in mouse bone marrow stromal ST2 cells and induction of alkaline phosphatase in mouse osteoblastic MC3T3-E1 cells. Notably, synthesis of SWNHs does not require a metal catalyst, the toxicity of which could become problematical in clinical use, and no cytotoxicity was observed in cells cultured in the presence of oxSWNHs under our conditions. Taken together, these observations highlight the potential utility of SWNHs in drug delivery systems.


Phase I studies of gemcitabine combined with carboplatin or paclitaxel.
Gemcitabine is a novel nucleoside analogue with a unique mechanism of action. In light of its good single-agent activity in several solid tumors, generally mild toxicity profile, and potential for synergy, combination phase I studies with other active chemotherapeutic agents have been conducted. In two studies the combination of gemcitabine and carboplatin was used to treat patients with non-small cell lung cancer. Gemcitabine was administered weekly x 3 every 4 weeks, and carboplatin was given on day 1. Although dose-limiting myelotoxicity was observed, encouraging activity was noted. In other studies patients with recurrent or persistent ovarian cancer or with refractory solid tumors were treated with weekly gemcitabine and paclitaxel on a 28-day schedule or with both drugs given every 2 weeks. Dose escalation was possible and toxicities were manageable. The effect of sequence of drug administration on the toxicity profile was also examined. Further trials to establish the efficacy of these promising approaches as well as combinations of all three drugs are needed.


The Association between Total Folate Intakes and Depression amongst Three Racial/Ethnic Groups.
Low dietary folate intake has been associated with depression outcomes, but few studies have been reported on the association in diverse populations. Using data from the National Health and Nutrition Examination Survey (NHANES), we examined the relationship between depression and folate intake from diet and supplementation in non-Hispanic whites, Hispanics and African Americans. 3,687 adult respondents from the 2009-2010 NHANES cycle were included. Statistical methods for analyzing data from complex survey sample designs were used to assess differences by race/ethnicity in demographic, behavioral, dietary and depression variables and to assess the relationship between depression and folate, adjusting for confounding variables using multivariable logistic regression. We observed significant (p < 0.01) differences by race/ethnicity for all demographic, behavioral, dietary and depression variables, except for physical activity. The relationship between dietary folate and depression significantly differed by race/ethnicity (p = 0.03), with an inverse and significant association in Hispanics only (OR= 0.25; 95% CI= 0.09 - 0.70.; p for trend = 0.02). These data suggest that a diet high in folate, such as from dark green leafy vegetables, may be associated with a reduced odds for depression, and specifically, Hispanics may benefit from nutrition education to potentially reduce depression in the population.


The rabbit as an infection model for equine proliferative enteropathy.
The objective of this study was to demonstrate the susceptibility of rabbits to Lawsonia intracellularis obtained from a case of clinical equine proliferative enteropathy (EPE). This is a preliminary step toward developing a rabbit infection model for studying pathogenesis and therapy of EPE in horses. Nine does were equally assigned to 3 groups. Animals in 2 groups (Group 1 and Group 2) were orally inoculated with different doses of cell-cultured L. intracellularis. Controls (Group 3) were sham-inoculated. Feces and blood were collected before the rabbits were infected and at 7, 14, and 21 days post-infection (DPI). Serum immunoglobulin G (IgG) titers were measured using an immunoperoxidase monolayer assay (IPMA) and fecal samples were analyzed with quantitative polymerase chain reaction (qPCR). A doe from each group was euthanized at 7, 14, and 21 DPI for collection and evaluation of intestinal samples. Tissues were stained by routine hematoxylin and eosin (H&E) method and immunohistochemistry (IHC) with L. intracellularis-specific mouse monoclonal antibody. At 14 DPI, serologic responses were detected in both infected groups, which maintained high titers through to 21 DPI. Lawsonia intracellularis DNA was detected in the feces of Group 2 on 7 DPI and in both infected groups on 14 DPI. Gross lesions were apparent in Group 1 and Group 2 on 14 DPI. Immunohistochemistry confirmed L. intracellularis antigen within cells of rabbits in Group 1 and Group 2 on 7, 14, and 21 DPI. No lesions, serologic response, shedding, or IHC labeling were found in Group 3 rabbits. This study describes an EPE rabbit model that simulates natural infection, as typical lesions, immune response, and fecal shedding were present.


Comparison of transcript profiles in late blight-challenged Solanum cajamarquense and B3C1 potato clones.
Two Solanum genotypes, a wild relative of cultivated potato S. cajamarquense (Cjm) and an advanced tetraploid clone B3C1 (B3), were inoculated with two Phytophthora infestans isolates and leaves were sampled at 72 and 96 h after inoculation. Gene expression in the inoculated versus noninoculated samples was monitored using the Institute of Genomic Research (TIGR) 10K potato array and real-time reverse transcriptase-polymerase chain reaction (RT-PCR). The current experiment is study number 83 of the TIGR expression profiling service project, and all data are publicly available in the Solanaceae Gene Expression Database (SGED) at ftp://ftp.tigr.org/pub/data/s_tuberosum/SGED. Differentially regulated cDNA clones were selected separately for each isolate-time point interaction by significant analysis of microarray (SAM), and differentially regulated clones were classified into functional categories by MapMan. The results show that the genes activated in B3 and Cjm have largely the same biological functions and are commonly activated when plants respond to pathogen attack. The genes activated within biological function categories were considerably different between the genotypes studied, suggesting that the defence pathways activated in B3 and Cjm during the tested conditions may involve unique genes. However, as indicated by real-time RT-PCR, some of the genes thought to be genotype specific may be activated across genotypes at other time points during disease development.


Radioprotective effects of genistein on HL-7702 cells via the inhibition of apoptosis and DNA damage.
Radiation induced normal tissue damage is the most important limitation for the delivery of a high potentially curative radiation dose. Genistein (GEN), one of the main soy isoflavone components, has drawn wide attention for its bioactivity in alleviating radiation damage. However, the effects and molecular mechanisms underlying the radioprotective effects of GEN remain unclear. In the present study, we showed that low concentration of GEN (1.5 µM) protected L-02 cells against radiation damage via inhibition of apoptosis, alleviation of DNA damage and chromosome aberration, down-regulation of GRP78 and up-regulation of HERP, HUS1 and hHR23A. In contrast, high concentration of GEN (20 µM) demonstrated radiosensitizing characteristics through the promotion of apoptosis and chromosome aberration, impairment of DNA repair, up-regulation of GRP78, and down-regulation of HUS1, SIRT1, RAD17, RAD51 and RNF8. These findings shed light on using low, but not high-concentration GEN, as a potential candidate for adjuvant therapy to alleviate radiation-induced injuries to human recipients of ionizing radiation.


Uptake, translocation, and metabolism of glyphosate, glufosinate, and dicamba mixtures in Echinochloa crus-galli and Amaranthus palmeri.
Echinochloa crus-galli (L.) Beauv. and Amaranthus palmeri S. Wats are two common and problematic weeds prevalent across the Midsouth of the USA. Herbicide absorption, translocation, and metabolism were investigated as potential sources of herbicide antagonism on A. palmeri and E. crus-galli using 14 C-labeled herbicides. Three 14 C-labeled herbicides, glyphosate, glufosinate, and dicamba, were utilized individually in separate experiments. Uptake of 14 C-glyphosate in E. crus-galli was 15% of the total applied radioactivity for glyphosate/glufosinate (897 + 595 g a.i./a.e. ha-1 ) compared to 25% for glyphosate alone. Similarly, uptake of 14 C-glyphosate in A. palmeri reduced by 10% when applied with glufosinate. Applying glyphosate/dicamba (897/560 g a.e. ha-1 ) reduced 14 C-glyphosate uptake in both species. In the 14 C-glufosinate experiment, both species absorbed less 14 C-glufosinate when mixed with glyphosate compared to glufosinate alone. No metabolic degradation of glyphosate was observed in either species. E. crus-galli metabolized dicamba 23 times faster than A. palmeri. When glufosinate was applied with dicamba, metabolic degradation of 14 C-dicamba was limited in both species. For example, 99.9% of the applied radioactivity was recovered in A. palmeri as the parent compound when 14 C-glufosinate dicamba was applied with glufosinate, compared to 95.7% for dicamba alone. These findings demonstrate absorption, translocation, or metabolism of dicamba, glufosinate, and glyphosate can be affected by mixing with another herbicide. As mixing two herbicides is often a critical component of resistance management, careful investigation into the performance of these mixtures in the field is needed. © 2020 Society of Chemical Industry.