Pseudohypoglycemia: a cause for unreliable finger-stick glucose measurements.
To identify patients with an inaccurate diagnosis of hypoglycemia and discuss predisposing factors. We describe our patient's clinical presentation, laboratory work-up, hospital course, and follow-up and review similar cases from the literature. A 27-year-old woman with Raynaud phenomenon was admitted because of symptomatic hypoglycemia. Physical examination showed tremulousness, sweating, and the classic Raynaud color changes of the hands during episodic symptoms. A 72-hour fast revealed finger-stick capillary glucose values ranging from 32 to 45 mg/dL on multiple occasions, while concurrent plasma glucose values were consistently 1.5 to 2 times higher. Capillary measurements of glucose performed in the arms and legs at room temperature and after warming of each extremity disclosed an increase in glucose levels from a range of 35 to 52 mg/dL at room temperature to a range of 82 to 100 mg/dL after warming, confirming a discordance between capillary and venous blood results. The diagnosis of pseudohypoglycemia was made. Pseudohypoglycemia has been reported in patients with Raynaud phenomenon, peripheral vascular disease, and shock and may result from increased glucose extraction by the tissues because of low capillary flow and increased glucose transit time. Pseudohypoglycemia should be suspected in the setting of impaired microcirculation and can be confirmed by readily available means.


On-line solid-phase preconcentration for sensitivity enhancement in capillary electrophoresis.
Capillary electrophoresis is attractive for the analysis of biological samples because only a few nanoliters of sample need to be injected. Indeed, optimal resolution is achieved when the injected volume is 1% or less of the total capillary volume. Unfortunately, this advantage leads to severe detection limitations compounded by the fact that many analytes in a biological sample are present at very low concentrations. To overcome the detection sensitivity limitations of CE, nonspecific, on-line preconcentration has been employed in a variety of applications. This technique is based on inserting a small quantity of reversed-phase material (e.g., C18 particles or membrane) near the inlet of the CE capillary for sample enrichment by solid-phase extraction. A detachable cartridge containing the solid phase is relatively simple to construct and permits the injection of large sample volumes (1-100 microL) into the capillary. Elution of adsorbed analyte in less than 100 nL of solvent permits a 100-1000 fold improvement in the effective concentration limit of detection, depending on the hydrophobicity of the compound. Detection of analytes present in complex mixtures at concentrations as low as 20 amol/microL has been reported when using an on-line preconcentration device in conjunction with CE-mass spectrometry. In this review, the potential merits of the technique are described and some examples of direct analysis of biological samples without rigorous off-line pretreatment are given.


Rich dynamics of a hepatitis B viral infection model with logistic hepatocyte growth.
Chronic hepatitis B virus (HBV) infection is a major cause of human suffering, and a number of mathematical models have examined within-host dynamics of the disease. Most previous HBV infection models have assumed that: (a) hepatocytes regenerate at a constant rate from a source outside the liver; and/or (b) the infection takes place via a mass action process. Assumption (a) contradicts experimental data showing that healthy hepatocytes proliferate at a rate that depends on current liver size relative to some equilibrium mass, while assumption (b) produces a problematic basic reproduction number. Here we replace the constant infusion of healthy hepatocytes with a logistic growth term and the mass action infection term by a standard incidence function; these modifications enrich the dynamics of a well-studied model of HBV pathogenesis. In particular, in addition to disease free and endemic steady states, the system also allows a stable periodic orbit and a steady state at the origin. Since the system is not differentiable at the origin, we use a ratio-dependent transformation to show that there is a region in parameter space where the origin is globally stable. When the basic reproduction number, R (0), is less than 1, the disease free steady state is stable. When R (0) > 1 the system can either converge to the chronic steady state, experience sustained oscillations, or approach the origin. We characterize parameter regions for all three situations, identify a Hopf and a homoclinic bifurcation point, and show how they depend on the basic reproduction number and the intrinsic growth rate of hepatocytes.


Herpes simplex virus type 1 VP22-mediated intercellular delivery of PTEN increases the antitumor activity of PTEN in esophageal squamous cell carcinoma cells in vitro and in vivo.
In the past decade, studies have revealed that the phosphatase and tensin homolog (PTEN) protein, a tumor suppressor, comprises a potential biological marker and therapeutic target for esophageal squamous cell carcinoma (ESCC). As such, the delivery of the PTEN gene represents a powerful strategy for ESCC therapy. The tegument protein VP22 of herpes simplex virus type 1 (HSV-1) has been reported to act as a transporter of heterologous proteins across the host cell membrane, thereby enhancing the biological functions of these proteins. In the present study, the intercellular delivery and antitumor activity of the fusion protein PTEN-VP22 were examined in the esophageal squamous cell carcinoma cell line Eca109 both in vitro and in vivo. VP22-mediated PTEN intercellular delivery was confirmed in the Eca109 cells by western blot analysis and by quantitation of immunofluorescence. VP22 alone did not exert antiproliferative effects or induce cell cycle arrest, induction of apoptosis, blockage of the Akt and focal adhesion kinase (FAK) pathways, tumor growth inhibition, or antiangiogenic effects in Eca109 cells. However, compared with PTEN alone, PTEN-VP22 exerted significantly higher antiproliferative effects and induced cell cycle arrest at G1 stage, apoptosis and antiangiogenic effects in Eca109 cells. Together, our findings demonstrate that VP22 alone does not exert antitumor activity directly; however, this protein mediates the intercellular delivery of PTEN and thereby increases its intracellular concentration to achieve a therapeutic steady state, leading to an overall increase in the antitumor activity of PTEN. This study provides further experimental data to confirm the potential of VP22-based intercellular delivery strategies for enhancing the efficacy of gene therapy for cancer treatment.


Efficient and robust prediction algorithms for protein complexes using Gomory-Hu trees.
Two-Hybrid (Y2H) Protein-Protein interaction (PPI) data suffer from high False Positive and False Negative rates, thus making searching for protein complexes in PPI networks a challenge. To overcome these limitations, we propose an efficient approach which measures connectivity between proteins not by edges, but by edge-disjoint paths. We model the number of edge-disjoint paths as a network flow and efficiently represent it in a Gomory-Hu tree. By manipulating the tree, we are able to isolate groups of nodes sharing more edge-disjoint paths with each other than with the rest of the network, which are our putative protein complexes. We examine the performance of our algorithm with Variation of Information and Separation measures and show that it belongs to a group of techniques which are robust against increased false positive and false negative rates. We apply our approach to yeast , mouse, worm, and human Y2H PPI networks, where it shows promising results. On yeast network, we identify 38 statistically significant protein clusters, 20 of which correspond to protein complexes and 16 to functional modules.


Penetration of topical indomethacin into phakic and aphakic rabbit eyes.
The penetration of indomethacin labeled with radioactive carbon (14C) into all ocular tissues and fluids was determined at various intervals in both phakic and aphakic rabbit eyes after either single or multiple (every 12 hours for three days) topical application (50 microL) regimens. More indomethacin was found in the vitreous of aphakic eyes compared with phakic eyes after single- or multiple-drop administration. Retinal and choroidal indomethacin concentrations were equal in both phakic and aphakic eyes after either drug regimen and are much greater than those of the vitreous. A pathway other than diffusion through the vitreous exists for the drug to reach these tissues. The concentration of indomethacin reaching the retina and choroid is not sufficient to inhibit prostaglandin formation locally. The concentrations of indomethacin are high enough to inhibit the biotransformation of prostaglandin precursor only in the anterior segment.


Allogeneic stem cell transplantation in second complete remission for core binding factor acute myeloid leukemia: a study from the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation.
Core binding factor acute myeloid leukemia (AML) comprises two subtypes with distinct cytogenetic abnormalities of either t(8;21)(q22;q22) or inv(16)(p13q22)/t(16;16)(p13;q22). Since long-term response to chemotherapy in these leukemias is relatively good, allogeneic hematopoietic stem cell transplantation is considered in patients who relapse and achieve second complete remission. To evaluate the outcomes of allogeneic transplantation in this indication, we studied 631 patients reported to the European Society for Blood and Marrow Transplantation Registry between the years 2000 and 2014. Leukemia-free survival probabilities at two and five years were 59.1% and 54.1%, while overall survival probabilities were 65% and 58.2%, respectively. The incidence of relapse and risk of non-relapse mortality at the same time points were 19.8% and 22.5% for relapse and 20.9% and 23.3% for non-relapse mortality, respectively. The most important adverse factors influencing leukemia-free and overall survival were: leukemia with t(8;21), presence of three or more additional chromosomal abnormalities, and Karnofsky performance score <80. Relapse risk was increased in t(8;21) leukemia and associated with additional cytogenetic abnormalities as well as reduced intensity conditioning. Measurable residual disease in molecular evaluation before transplantation was associated with increased risk of relapse and inferior leukemia-free survival.


JAtlasView: a Java atlas-viewer for browsing biomedical 3D images and atlases.
Many three-dimensional (3D) images are routinely collected in biomedical research and a number of digital atlases with associated anatomical and other information have been published. A number of tools are available for viewing this data ranging from commercial visualization packages to freely available, typically system architecture dependent, solutions. Here we discuss an atlas viewer implemented to run on any workstation using the architecture neutral Java programming language. We report the development of a freely available Java based viewer for 3D image data, descibe the structure and functionality of the viewer and how automated tools can be developed to manage the Java Native Interface code. The viewer allows arbitrary re-sectioning of the data and interactive browsing through the volume. With appropriately formatted data, for example as provided for the Electronic Atlas of the Developing Human Brain, a 3D surface view and anatomical browsing is available. The interface is developed in Java with Java3D providing the 3D rendering. For efficiency the image data is manipulated using the Woolz image-processing library provided as a dynamically linked module for each machine architecture. We conclude that Java provides an appropriate environment for efficient development of these tools and techniques exist to allow computationally efficient image-processing libraries to be integrated relatively easily.


Spontaneous regression of a pulmonary adenocarcinoma after core needle biopsy.
Spontaneous regression (SR) of cancer, especially lung cancer, is a rare biological event with a mechanism that is not currently understood. Immunological mechanisms seem to be the stronger explanation in SR of a lung cancer. We report the rare case of SR of a lung adenocarcinoma stage IA, in a 75-year-old man, which was incidentally diagnosed and histologically confirmed. Due to the patient's comorbidities and his poor pulmonary function, stereotactic radiotherapy was scheduled. However, by the time the treatment was due to start, the tumor was no longer detectable.


Selective inhibition of interleukin-4 gene expression in human T cells by aspirin.
Previous studies indicated that aspirin (acetylsalicylic acid [ASA]) can have profound immunomodulatory effects by regulating cytokine gene expression in several types of cells. This study is the first in which concentrations of ASA in the therapeutic range were found to significantly reduce interleukin (IL)-4 secretion and RNA expression in freshly isolated and mitogen-primed human CD4+ T cells. In contrast, ASA did not affect IL-13, interferon-gamma, and IL-2 expression. ASA inhibited IL-4, but not IL-2, promoter-driven chloramphenicol acetyltransferase expression in transiently transfected Jurkat T cells. The structurally unrelated nonsteroidal anti-inflammatory drugs indomethacin and flurbiprofen did not affect cytokine gene expression in T cells, whereas the weak cyclo-oxygenase inhibitor salicylic acid was at least as effective as ASA in inhibiting IL-4 expression and promoter activity. The inhibitory effect of ASA on IL-4 transcription was not mediated by decreased nuclear expression of the known salicylate target nuclear factor (NF)-kappaB and was accompanied by reduced binding of an inducible factor to an IL-4 promoter region upstream of, but not overlapping, the NF of activated T cells- and NF-kappaB-binding P1 element. It is concluded that anti-inflammatory salicylates, by means of a previously unrecognized mechanism of action, can influence the nature of adaptive immune responses by selectively inhibiting the expression of IL-4, a critical effector of these responses, in CD4+ T cells.


Predictability before transplant of hepatic complications following allogeneic bone marrow transplantation.
This study was undertaken to evaluate the occurrence of VOD and other liver diseases following BMT in a patient population with a high incidence of hepatitis before conditioning regimen. We prospectively reviewed 186 consecutive patients undergoing BMT from 1976 to 1986 to determine incidence and type of liver disease after BMT and predisposing factors. Two of 186 patients experienced VOD (1.07%). Acute and chronic liver GVHD were found in 25.8% and 36% of the patients, respectively. Acute hepatitis (AH) was diagnosed in 29.4% and chronic hepatitis (CH) in 42.6% of the patients. Statistical analysis showed no influence of pretransplant variables on the occurrence of acute GVHD and AH; there was a weak correlation (P = 0.01) between pre-BMT abnormal transaminases and occurrence of chronic GVHD. Contingency table and Cox analysis showed a greater risk of CH for patients with abnormal pretransplant SGPT levels (P = 0.0004 and P = 0.0022). No other variables could be associated with posttransplant CH. Actuarial survival was 71% versus 69% for patients with normal versus abnormal transaminases (P = 0.2). As VOD was a rare event, despite 53% of patients having abnormal transaminase values before transplant, we suggest that a lower and slower TBI is more important than pretransplant normal transaminases in preventing this complication. We conclude that evidence of compensated hepatitis is not a relative contraindication for BMT.


Thread-Like Radical-Polymerization via Autonomously Propelled (TRAP) Bots.
Micromotor-mediated synthesis of thread-like hydrogel microstructures in an aqueous environment is presented. The study utilizes a catalytic micromotor assembly (owing to the presence of a Pt layer), with an on-board chemical reservoir (i.e., polymerization mixture), toward thread-like radical-polymerization via autonomously propelled bots (i.e., TRAP bots). Synergistic coupling of catalytically active Pt layer, together with radical initiators (H2 O2 and FeCl3 (III)), and PEGDA monomers preloaded into the TRAP bot, results in the polymerization of monomeric units into elongated thread-like hydrogel polymers coupled with self-propulsion. Interestingly, polymer generation via TRAP bots can also be triggered in the absence of hydrogen peroxide for cellular/biomedical application. The resulting polymeric hydrogel microstructures are able to entrap living cells (NIH 3T3 fibroblast cells), and are easily separable via a centrifugation or magnetic separation (owing to the presence of a Ni layer). The cellular biocompatibility of TRAP bots is established via a LIVE/DEAD assay and MTS cell proliferation assay (7 days observation). This is the first study demonstrating real-time in situ hydrogel polymerization via an artificial microswimmer, capable of enmeshing biotic/abiotic microobjects in its reaction environment, and lays a strong foundation for advanced applications in cell/tissue engineering, drug delivery, and cleaner technologies.


Appetite testing in HIV-infected African adults recovering from malnutrition and given antiretroviral therapy.
The Nutritional Support for Africans Starting Antiretroviral Therapy (NUSTART) trial was designed to determine whether nutritional support for malnourished HIV-infected adults starting antiretroviral therapy (ART) can improve early survival. Appetite is related to health outcomes in this population, but the optimal appetite metric for field use is uncertain. We evaluated two measures of appetite with the goal of improving understanding and treatment of malnutrition in HIV-infected adults. Longitudinal cohort study embedded in a clinical trial of vitamin and mineral-fortified, v. unfortified, lipid-based nutritional supplements. HIV clinics in Mwanza, Tanzania and Lusaka, Zambia. Malnourished (BMI<18.5 kg/m2) HIV-infected adults starting ART. Appetite measurements, by short questionnaire and by weight of maize porridge consumed in a standardized test, were compared across time and correlated with changes in weight. Appetite questionnaire scores, from polychoric correlation, and porridge test results were normally distributed for Tanzanians (n 187) but clustered and unreliable for Zambians (n 297). Among Tanzanian patients, the appetite score increased rapidly from referral for ART, plateaued at the start of ART and then increased slowly during the 12-week follow-up. Change in appetite questionnaire score, but not porridge test, correlated with weight change in the corresponding two-week intervals (P=0.002) or over the whole study (P=0.05) but a point estimate of hunger did not predict weight change (P=0.4). In Tanzania change in appetite score correlated with weight change, but single point measurements did not. Appetite increases several weeks after the start of ART, which may be an appropriate time for nutritional interventions for malnourished HIV-infected adults.


Claims for the anabolic effects of growth hormone: a case of the emperor's new clothes?
This review examines the evidence that growth hormone has metabolic effects in adult human beings. The conclusion is that growth hormone does indeed have powerful effects on fat and carbohydrate metabolism, and in particular promotes the metabolic use of adipose tissue triacylglycerol. However, there is no proof that net protein retention is promoted in adults, except possibly of connective tissue. The overexaggeration of the effects of growth hormone in muscle building is effectively promoting its abuse and thereby encouraging athletes and elderly men to expose themselves to increased risk of disease for little benefit.


Vitamin D, the placenta and early pregnancy: effects on trophoblast function.
Pregnancy is associated with significant changes in vitamin D metabolism, notably increased maternal serum levels of active vitamin D, 1,25-dihydroxyvitamin (1,25(OH)2D). This appears to be due primarily to increased renal activity of the enzyme 25-hydroxyvitamin D-1α-hydroxylase (CYP27B1) that catalyzes synthesis of 1,25(OH)2D, but CYP27B1 expression is also prominent in both the maternal decidua and fetal trophoblast components of the placenta. The precise function of placental synthesis of 1,25(OH)2D remains unclear, but is likely to involve localized tissue-specific responses with both decidua and trophoblast also expressing the vitamin D receptor (VDR) for 1,25(OH)2D. We have previously described immunomodulatory responses to 1,25(OH)2D by diverse populations of VDR-expressing cells within the decidua. The aim of the current review is to detail the role of vitamin D in pregnancy from a trophoblast perspective, with particular emphasis on the potential role of 1,25(OH)2D as a regulator of trophoblast invasion in early pregnancy. Vitamin D deficiency is common in pregnant women, and a wide range of studies have linked low vitamin D status to adverse events in pregnancy. To date, most of these studies have focused on adverse events later in pregnancy, but the current review will explore the potential impact of vitamin D on early pregnancy, and how this may influence implantation and miscarriage.


A central role for CD68(+) macrophages in hepatopulmonary syndrome. Reversal by macrophage depletion.
The etiology of hepatopulmonary syndrome (HPS), a common complication of cirrhosis, is unknown. Inflammation and macrophage accumulation occur in HPS; however, their importance is unclear. Common bile duct ligation (CBDL) creates an accepted model of HPS, allowing us to investigate the cause of HPS. We hypothesized that macrophages are central to HPS and investigated the therapeutic potential of macrophage depletion. Hemodynamics, alveolar-arterial gradient, vascular reactivity, and histology were assessed in CBDL versus sham rats (n = 21 per group). The effects of plasma on smooth muscle cell proliferation and endothelial tube formation were measured. Macrophage depletion was used to prevent (gadolinium) or regress (clodronate) HPS. CD68(+) macrophages and capillary density were measured in the lungs of patients with cirrhosis versus control patients (n = 10 per group). CBDL increased cardiac output and alveolar-arterial gradient by causing capillary dilatation and arteriovenous malformations. Activated CD68(+)macrophages (nuclear factor-κB+) accumulated in HPS pulmonary arteries, drawn by elevated levels of plasma endotoxin and lung monocyte chemoattractant protein-1. These macrophages expressed inducible nitric oxide synthase, vascular endothelial growth factor, and platelet-derived growth factor. HPS plasma increased endothelial tube formation and pulmonary artery smooth muscle cell proliferation. Macrophage depletion prevented and reversed the histological and hemodynamic features of HPS. CBDL lungs demonstrated increased medial thickness and obstruction of small pulmonary arteries. Nitric oxide synthase inhibition unmasked exaggerated pulmonary vasoconstrictor responses in HPS. Patients with cirrhosis had increased pulmonary intravascular macrophage accumulation and capillary density. HPS results from intravascular accumulation of CD68(+)macrophages. An occult proliferative vasculopathy may explain the occasional transition to portopulmonary hypertension. Macrophage depletion may have therapeutic potential in HPS.


Serum antibodies specific to CD outer membrane protein of Moraxella catarrhalis, P6 outer membrane protein of non-typeable Haemophilus influenzae and capsular polysaccharides of Streptococcus pneumoniae in children with otitis media with effusion.
We measured the levels of serum IgG antibodies to CD outer membrane protein of Moraxella catarrhalis, P6 outer membrane protein of non-typeable Haemophilus influenzae and capsular polysaccharides of Streptococcus pneumoniae in 168 children with otitis media with effusion (OME) who were followed prospectively, using ELISA. Serum IgG antibodies to CD, P6 and pneumococcal capsular polysaccharides were detected in all samples. The anti-pneumococcal polysaccharides antibody level was highest, followed by the anti-P6 antibody level and anti-CD antibody was lowest (median:interquartile ranges were 45.9:19.1-100 microg/ml, 15.6:9.70-23.2 microg/ml and 1.06:0.73-1.87 microg/ml, respectively). In children aged 0-6 years, there were positive correlations among the antibody levels (anti-CD vs anti-P6, r=0.325, p <0.001; anti-CD vs anti-polysaccharide, r=0.397, p <0.0001; anti-P6 vs anti-polysaccharide, r=0.175, p=0.057). However, no relationship was seen in children aged 7-15 years. Children were classified according to severity of OME during the 1-year follow-up. In children aged 0-6 years, the severity of OME correlated inversely with the levels of anti-CD antibody (r=-.23, p=0.012), of anti-P6 antibody (r=-0.292, p=0.0015), and of anti-pneumococcal polysaccharides antibody (r=-0.25, p=0.0064). However, no correlation was found between antibody levels and severity of OME in children aged 7-15 years. These data suggest that persistence and/or recurrence of OME may be due to an insufficient serum antibody response to middle ear pathogens in young children.


A novel hemolytic complement-sufficient NSG mouse model supports studies of complement-mediated antitumor activity in vivo.
Monoclonal antibodies (mAbs) have emerged as a mainstream therapeutic option against cancer. mAbs mediate tumor cell-killing through several mechanisms including complement-dependent cytotoxicity (CDC). However, studies of mAb-mediated CDC against tumor cells remain largely dependent on in vitro systems. Previously developed and widely used NOD-scid IL2rγnull (NSG) mice support enhanced engraftment of many primary human tumors. However, NSG mice have a 2-bp deletion in the coding region of the hemolytic complement (Hc) gene, and it is not possible to evaluate CDC activity in NSG mice. To address this limitation, we generated a novel strain of NSG mice-NSG-Hc1-that have an intact complement system able to generate the membrane attack complex. Utilizing the Daudi Burkitt's human lymphoma cell line, and the anti-human CD20 mAb rituximab, we further demonstrated that the complement system in NSG-Hc1 mice is fully functional. NSG-Hc1 mice expressed CDC activity against Daudi cells in vivo following rituximab treatment and showed longer overall survival compared with rituximab-treated NSG mice that lack hemolytic complement. Our results validate the NSG-Hc1 mouse model as a platform for testing mechanisms underlying CDC in vivo and suggest its potential use to compare complement-dependent and complement-independent cytotoxic activity mediated by therapeutic mAbs.


Rat parotid gland cell differentiation in three-dimensional culture.
The use of polarized salivary gland cell monolayers has contributed to our understanding of salivary gland physiology. However, these cell models are not representative of glandular epithelium in vivo, and, therefore, are not ideal for investigating salivary epithelial functions. The current study has developed a three-dimensional (3D) cell culture model for rat Par-C10 parotid gland cells that forms differentiated acinar-like spheres on Matrigel. These 3D Par-C10 acinar-like spheres display characteristics similar to differentiated acini in salivary glands, including cell polarization, tight junction (TJ) formation required to maintain transepithelial potential difference, basolateral expression of aquaporin-3 and Na+/K+/2Cl- cotransporter-1, and responsiveness to the muscarinic receptor agonist carbachol that is decreased by the anion channel blocker diphenylamine-2-carboxylic acid or chloride replacement with gluconate. Incubation of the spheres in the hypertonic medium increased the expression level of the water channel aquaporin-5. Further, the proinflammatory cytokines tumor necrosis factor-alpha and interferon-gamma induced alterations in TJ integrity in the acinar-like spheres without affecting individual cell viability, suggesting that cytokines may affect salivary gland function by altering TJ integrity. Thus, 3D Par-C10 acinar-like spheres represent a novel in vitro model to study physiological and pathophysiological functions of differentiated acini.


Patterns of GABA(B)R1a,b receptor gene expression in monkey and human visual cortex.
In situ hybridization histochemistry and immunocytochemistry were used to examine GABA(B)R1a,b receptor mRNA and protein expression in areas 17 and 18 of the visual cortex of normal macaque monkeys and of monkeys that had been deprived of vision in one eye. In addition, the normal patterns of GABA(B)R1a,b protein expression were immunocytochemically studied in the human visual cortex. Overall levels of GABA(B)R1a,b transcript were higher in area 17 than in area 18. In area 17 GABA(B)R1a,b mRNA levels were highest in layers IVC and VI, moderate in layers II-IVA and low in layers I, IVB and V. In area 18 GABA(B)R1a,b transcript expression was high in layers II and III, moderate in layers IV and VI and low in layers I and V. Immunocytochemistry revealed nearly identical patterns of GABA(B)R1a,b protein expression in areas 17 and 18 in monkey and human. Both pyramidal and non-pyramidal neurons were GABA(B)R1a,b immunoreactive. The majority of intensely immunoreactive neurons in layers II, III, V and VI were pyramidal cells. Numerous non-pyramidal cells were intensely immunoreactive in layer IV of area 17 but layer IV cells were only lightly immunoreactive in area 18. Following 10 day periods of monocular deprivation, induced by intravitreal injections of tetrodotoxin, levels of GABA(B)R1a,b mRNA and protein were decreased in the deprived eye dominance columns of layers IVC and VI.


A simple rectification method for linear multi-baseline stereovision system.
The linear multi-baseline stereo system introduced by the CMU-RI group has been proven to be a very effective and robust stereovision system. However, most traditional stereo rectification algorithms are all designed for binocular stereovision system, and so, cannot be applied to a linear multi-baseline system. This paper presents a simple and intuitional method that can simultaneously rectify all the cameras in a linear multi-baseline system. Instead of using the general 8-parameter homography transform, a two-step virtual rotation method is applied for rectification, which results in a more specific transform that has only 3 parameters, and more stability. Experimental results for real stereo images showed the presented method is efficient.


Analyzing mixing systems using a new generation of Bayesian tracer mixing models.
The ongoing evolution of tracer mixing models has resulted in a confusing array of software tools that differ in terms of data inputs, model assumptions, and associated analytic products. Here we introduce MixSIAR, an inclusive, rich, and flexible Bayesian tracer (e.g., stable isotope) mixing model framework implemented as an open-source R package. Using MixSIAR as a foundation, we provide guidance for the implementation of mixing model analyses. We begin by outlining the practical differences between mixture data error structure formulations and relate these error structures to common mixing model study designs in ecology. Because Bayesian mixing models afford the option to specify informative priors on source proportion contributions, we outline methods for establishing prior distributions and discuss the influence of prior specification on model outputs. We also discuss the options available for source data inputs (raw data versus summary statistics) and provide guidance for combining sources. We then describe a key advantage of MixSIAR over previous mixing model software-the ability to include fixed and random effects as covariates explaining variability in mixture proportions and calculate relative support for multiple models via information criteria. We present a case study of Alligator mississippiensis diet partitioning to demonstrate the power of this approach. Finally, we conclude with a discussion of limitations to mixing model applications. Through MixSIAR, we have consolidated the disparate array of mixing model tools into a single platform, diversified the set of available parameterizations, and provided developers a platform upon which to continue improving mixing model analyses in the future.


Electroacupuncture Treatment Attenuates Paclitaxel-Induced Neuropathic Pain in Rats via Inhibiting Spinal Glia and the TLR4/NF-κB Pathway.
Neuropathic pain is a major side-effect of paclitaxel (PTX) chemotherapy. Although the precise mechanisms responsible for this pain are unclear, the activation of neuroglia and upregulation of the TLR4/NF-κB pathway are known to be involved. In this study, we determined whether electroacupuncture (EA) could limit mechanical hypersensitivity resulting from the chemotherapeutic drug PTX in rats, and investigated the potential mechanisms involved. Rats intraperitoneally received a cumulative dose of 8 mg/kg PTX (2 mg/kg per day) or vehicle control on alternate days (day 0, 2, 4 and 6). EA treatment (10 Hz, 1 mA) was applied at bilateral ST36 acupoints in rats once every other day on days 0-14. For sham EA, needles were inserted at ST36 acupoints without electrical stimulation. Mechanical allodynia was measured by mechanical withdrawal latency (MWL) of paws to a mechanical stimulus every 2 days. Protein expression of TLR4 and NF-κB p65, as well as TMEM119 and GFAP (indicators of microglia and astrocytes, respectively) in spinal cord was quantified by Western blot analysis. Levels of inflammatory cytokines IL-1β and TNF-α in spinal cord and serum were detected by ELISA. Mechanical allodynia induced by PTX in both paws (right and left) of rats was significantly attenuated by EA but not sham EA treatment. In addition, EA, but not sham EA, inhibited the activation of both microglia (TMEM119) and astrocytes (GFAP) in lumbar spinal cord. Moreover, Western blot analysis revealed that protein expression of TLR4 and NF-κB in spinal cord was suppressed by EA but not sham EA treatment. PTX significantly increased inflammatory cytokines in spinal cord and serum, which were ameliorated by EA treatment but not by sham EA. These results indicate that EA treatment attenuates PTX-induced mechanical allodynia. The putative mechanism corroborating this finding could be related to the suppression of activated microglia and astrocytes in spinal cord, as well as the inhibition of the activated TLR4/NF-κB signaling pathway by EA treatment.


Time to analgesia for patients with painful extremity injuries transported to the emergency department by ambulance.
To measure the time to analgesia for patients with painful, isolated extremity injuries brought to the emergency department (ED) by emergency medical services (EMS). A retrospective chart review of all patients presenting with isolated, painful extremity injuries during an 18-month period to a Level 1 trauma center. Medical records were reviewed by diagnostic codes for extremity injuries. patients 18 years or older transported to the ED by EMS; isolated, painful extremity injuries; received parenteral analgesia in the ED or by EMS. Excluded: multiple trauma patients, interfacility transfers, hemodynamically unstable patients, head-injured patients, intoxicated patients, and patients with mental status changes. Data elements: age, sex, EMS arrival time, EMS medication time, hospital triage time, and ED medication time. All "times to analgesia" were calculated from the EMS arrival time on scene. Extremity injuries were identified in 706 patients. Of these, 104 patients with painful, isolated, extremity injuries met all inclusion criteria. Thirteen (12.5%) of 104 received analgesia by EMS during prehospital care. Ninety-one patients (88%) first received parenteral analgesia in the ED. The mean time to analgesia for EMS treated patients was 23 minutes (95% CI 16.7-30.2). Mean time to analgesia for patients treated in the ED was 113 minutes (95% CI 99.2-128.1). Mean time to analgesia after triage in this group was 75 minutes (95% CI 60.8-89.7). In this study, patients received analgesia sooner when administered by EMS during prehospital care. There was a significant time delay after triage for patients first medicated in the ED.


Family-Based Behavioral Treatment for Childhood Obesity Implemented in Pediatric Primary Care: A Randomized Clinical Trial.
Intensive behavioral interventions for childhood overweight and obesity are recommended by national guidelines, but are currently offered primarily in specialty clinics. Evidence is lacking on their effectiveness in pediatric primary care settings. To evaluate the effects of family-based treatment for overweight or obesity implemented in pediatric primary care on children and their parents and siblings. This randomized clinical trial in 4 US settings enrolled 452 children aged 6 to 12 years with overweight or obesity, their parents, and 106 siblings. Participants were assigned to undergo family-based treatment or usual care and were followed up for 24 months. The trial was conducted from November 2017 through August 2021. Family-based treatment used a variety of behavioral techniques to develop healthy eating, physical activity, and parenting behaviors within families. The treatment goal was 26 sessions over a 24-month period with a coach trained in behavior change methods; the number of sessions was individualized based on family progress. The primary outcome was the child's change from baseline to 24 months in the percentage above the median body mass index (BMI) in the general US population normalized for age and sex. Secondary outcomes were the changes in this measure for siblings and in BMI for parents. Among 452 enrolled child-parent dyads, 226 were randomized to undergo family-based treatment and 226 to undergo usual care (child mean [SD] age, 9.8 [1.9] years; 53% female; mean percentage above median BMI, 59.4% [n = 27.0]; 153 [27.2%] were Black and 258 [57.1%] were White); 106 siblings were included. At 24 months, children receiving family-based treatment had better weight outcomes than those receiving usual care based on the difference in change in percentage above median BMI (-6.21% [95% CI, -10.14% to -2.29%]). Longitudinal growth models found that children, parents, and siblings undergoing family-based treatment all had outcomes superior to usual care that were evident at 6 months and maintained through 24 months (0- to 24-month changes in percentage above median BMI for family-based treatment and usual care were 0.00% [95% CI, -2.20% to 2.20%] vs 6.48% [95% CI, 4.35%-8.61%] for children; -1.05% [95% CI, -3.79% to 1.69%] vs 2.92% [95% CI, 0.58%-5.26%] for parents; and 0.03% [95% CI, -3.03% to 3.10%] vs 5.35% [95% CI, 2.70%-8.00%] for siblings). Family-based treatment for childhood overweight and obesity was successfully implemented in pediatric primary care settings and led to improved weight outcomes over 24 months for children and parents. Siblings who were not directly treated also had improved weight outcomes, suggesting that this treatment may offer a novel approach for families with multiple children. ClinicalTrials.gov Identifier: NCT02873715.


Sex-Biased Aging Effects on Ig Somatic Hypermutation Targeting.
Aged individuals, particularly males, display an impaired level of Ab response compared with their younger counterparts, yet the molecular mechanisms responsible for the discrepancy are not well understood. We hypothesize that some of this difference may be linked to B cell somatic hypermutation (SHM) targeting, including error-prone DNA repair activities that are crucial to Ab diversification. To examine the effects of aging on SHM targeting, we analyzed B cell Ig repertoire sequences from 27 healthy male and female human subjects aged 20-89. By studying mutation patterns based on 985,069 mutations obtained from 123,415 sequences, we found that the SHM mutability hierarchies on microsequence motifs (i.e., SHM hot/cold spots) are mostly consistent between different age and sex groups. However, we observed a lower frequency in mutations involving Phase II SHM DNA repair activities in older males, but not in females. We also observed, from a separate study, a decreased expression level of DNA mismatch repair genes involved in SHM in older individuals compared with younger individuals, with larger fold changes in males than in females. Finally, we showed that the balance between Phase I versus Phase II SHM activities impacts the resulting Ig phenotypes. Our results showed that the SHM process is altered in some older individuals, providing insights into observed clinical differences in immunologic responses between different age and sex groups.


Synthesis and antiviral activity of 5‑(4‑chlorophenyl)-1,3,4-thiadiazole sulfonamides.
Starting from 4-chlorobenzoic acid, 10 new 5-(4-chlorophenyl)-N-substituted-N-1,3,4-thiadiazole-2-sulfonamide derivatives were synthesized in six-steps. Esterification of 4-chlorobenzoic acid with methanol and subsequent hydrazination, salt formation and cyclization afforded 5-(4-chlorophen-yl)-1,3,4-thiadiazole-2-thiol (5). Conversion of this intermediate into sulfonyl chloride 6, followed by nucleophilic attack of the amines gave the title sulfonamides 7a-7j whose structures were confirmed by NMR, IR and elemental analysis. The bioassay tests showed that compounds 7b and 7i possessed certain anti-tobacco mosaic virus activity.


Homology of Pseudomonas cytochrome c-551 with eukaryotic c-cytochromes.
The homology of Pseudomonas cytochrome c-551 with eukaryotic cytochromes c is examined with a computer-based procedure devised to determine whether similarities exist between these proteins. One method is given by which the more recently evolved cytochromes c might have arisen from the Pseudomonas protein. This procedure involves only common genetic phenomena and accounts for most of the structural differences between the bacterial and mammalian cytochromes. A time-scale relationship between the c-cytochromes from several microorganisms, a mold, yeast, and the eukaryotic organisms is proposed.


Neotropical ant gardens II. Bioassays of seed compounds.
A number of volatile compounds occur on the seeds of taxonomically unrelated ant-garden epiphytes in western Amazonia. In field trials in southeastern Peru, we assayed the responses of ant-garden ants (Camponotus femoratus) to these and structurally similar compounds applied to artificial "seeds" made from zeolite molecular sieves. Benzothiazole,2, present on seeds of eight ant-garden epiphytes, repelled ants over the range of concentrations tested, as did 1-(2-hydroxy-6-methylphenyl)ethanone,3, occurring on seeds of six ant-garden epiphytes. 2-Hydroxy-6-methylbenzoic acid, methyl ester (methyl-6-methylsalicylate, 6-MMS),1, found on seeds of at least nine ant-garden epiphytes, was mildly repellent at high concentration, but stimulated excitement, seed handling, and (rarely) seed carrying at lower concentrations. Vanillin,5, a seed compound of four ant-garden epiphytes, and limonene,6, a monoterpene from seeds of three ant-garden epiphytes, both stimulated excitement, alarm, seed handling, and (rarely) seed carrying. Identified from seeds of seven ant-garden epiphytes, 1-(2,4-dihydroxyphenyl)ethanone,4, elicited little or no response. Among 70 compounds tested (mainly aromatic compounds), those found on seeds of ant-garden epiphytes or having structural features in common with such compounds were the most attractive to the ants. Although not present on epiphyte seeds, 2-hydroxy-3-methoxybenzenemethanol,10, consistently stimulated seed transport to the nest in one year, but did so only rarely in subsequent years. Some of the volatile compounds on seeds of ant-garden epiphytes probably play a role in ant attraction to epiphyte seeds, but evidence remains ambiguous. Finally,Ca. femoratus responded to one test compound [1-(2-hydroxy-4-methoxyphenyl)ethanone,60] (absent from epiphyte seeds) by descending from the vegetation to the ground.


Urgency and other lower urinary tract symptoms in men aged ≥ 40 years: a Belgian epidemiological survey using the ICIQ-MLUTS questionnaire.
The aim of this study was to assess prevalence and associated bother of male lower urinary tract symptoms (LUTS), especially of urgency and other storage symptoms, since these are generally assumed to be underdiagnosed/undertreated in men. Data on bladder condition of men ≥ 40 years were prospectively collected by 124 general practitioners (GPs) during a regular visit for any reason, using the validated ICIQ-MLUTS questionnaire. For 13 symptoms, prevalence (scale 0-4) and bother (scale 0-10) were scored. General bladder-related quality of life (scale 0-3) was also assessed. Data from 5890 men (mean age: 61.2 years) were analysed. A total of 7.7% had urgency most or all of the time (score ≥ 3) and 6.2% had bothersome urgency (score ≥ 3 + bother score ≥ 5). Nocturia (69.2%) and urgency (58.3%) were the most prevalent and bothersome symptoms. Both prevalence and bother of all LUTS increased with age. Additionally, 28.9% of men reported to be a little bothered by their bladder condition in everyday life, while 11.9% were bothered a lot/very much (2.5% in age group 40-49 years increasing to 29.2% in those > 80 years). In the general population of men ≥ 40 years who visited a GP for any reason, 41% indicated to be at least a little bothered by their bladder condition. The prevalence of LUTS, especially nocturia and urgency, is high and a significant number of men indicated to be seriously bothered. Increasing awareness of male LUTS, and storage symptoms in particular, is warranted to discuss management options that could increase quality of life.


The role of hydrogen peroxide and nitric oxide in the induction of plant-encoded RNA-dependent RNA polymerase 1 in the basal defense against Tobacco mosaic virus.
Plant RNA-dependent RNA Polymerase 1 (RDR1) is an important element of the RNA silencing pathway in the plant defense against viruses. RDR1 expression can be elicited by viral infection and salicylic acid (SA), but the mechanisms of signaling during this process remains undefined. The involvement of hydrogen peroxide (H2O2) and nitric oxide (NO) in RDR1 induction in the compatible interactions between Tobacco mosaic tobamovirus (TMV) and Nicotiana tabacum, Nicotiana benthamiana, and Arabidopsis thaliana was examined. TMV inoculation onto the lower leaves of N. tabacum induced the rapid accumulation of H2O2 and NO followed by the increased accumulation of RDR1 transcripts in the non-inoculated upper leaves. Pretreatment with exogenous H2O2 and NO on upper leaf led to increased RDR1 expression and systemic TMV resistance. Conversely, dimethylthiourea (an H2O2 scavenger) and 2-(4-carboxyphenyl)- 4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (an NO scavenger) partly blocked TMV- and SA-induced RDR1 expression and increased TMV susceptibility, whereas pretreatment with exogenous H2O2 and NO failed to diminish TMV infection in N. benthamiana plants with naturally occurring RDR1 loss-of-function. Furthermore, in N. tabacum and A. thaliana, TMV-induced H2O2 accumulation was NO-dependent, whereas NO generation was not affected by H2O2. These results suggest that, in response to TMV infection, H2O2 acts downstream of NO to mediate induction of RDR1, which plays a critical role in strengthening RNA silencing to restrict systemic viral infection.


CREB controls LAP/C/EBP beta transcription.
LAP/C/EBP beta is a member of the C/EBP family of transcription factors and is involved in hepatocyte-specific gene expression. Recently we showed that, besides its posttranscriptional regulation, LAP/C/EBP beta mRNA is modulated during liver regeneration. Therefore, in this study we investigated mechanisms which control LAP/C/EBP beta gene transcription. Deletion analysis of the 5'-flanking region, located upstream of the start site of transcription in the LAP/C/EBP beta gene, demonstrated that a small region in close proximity to the TATA box is important in maintaining a high level of transcription of the luciferase reporter gene constructs. In gel shift experiments two sites were identified which are important for specific complex formation within this region. Further analysis by cross-linking, super shift, and competition experiments was performed with liver cell nuclear extracts, hepatoma cell nuclear extracts, or recombinant CREB protein. These experiments conclusively demonstrated that CREB binds to both sites in the LAP/C/EBP beta promoter with an affinity similar to that with the CREB consensus sequence. Transfection experiments with promoter constructs where the CREB sites were mutated showed that these sites are important to maintain both basal promoter activity and LAP/C/EBP beta inducibility through CREB. Northern blot analysis and runoff transcription assays demonstrated that the protein kinase A pathway not only stimulated the activity of the luciferase reporter construct but also the transcription of the endogenous LAP/C/EBP beta gene in different cell types. Western blot analysis of rat liver cell nuclear extracts and runoff transcription assays of rat liver cell nuclei after two-thirds hepatectomy showed a functional link between the induction of CREB phosphorylation and LAP/C/EBP beta mRNA transcription during liver regeneration. These results demonstrate that the two CREB sites are important to control LAP/C/EBP beta transcription in vivo. As several pathways control CREB phosphorylation, our results provide evidence for the transcriptional regulation of LAP/C/EBP beta via CREB under different physiological conditions.


Abnormal glucose metabolism among older men with or at risk of HIV infection.
To determine factors associated with diabetes, insulin resistance, and abnormal glucose tolerance in older men with or at risk of HIV infection. Diabetes was assessed by self-report in 643 men >or=49 years old with or at risk of HIV infection. In a subset of 216 men without previously diagnosed diabetes [including 90 HIV-uninfected men, 28 HIV-infected, antiretroviral-naive men, 28 HIV-infected men taking non-protease inhibitor (PI)-containing highly active antiretroviral therapy (HAART), and 70 HIV-infected men taking PI-containing HAART], an oral glucose tolerance test with insulin levels was performed. HIV serology, CD4 cell count, weight, height and waist circumference were measured. Antiretroviral use, drug use, family history of diabetes, physical activity and sociodemographic data were obtained using standardized interviews. Of 643 participants, 116 (18%) had previously diagnosed diabetes. With the oral glucose tolerance test, 15 of 216 men (7%) were found to have undiagnosed diabetes and 40 (18%) impaired glucose tolerance. Factors independently associated with previously diagnosed diabetes included use of non-PI-containing HAART, methadone treatment, positive CAGE test for alcoholism, obesity and family history of diabetes. Factors independently associated with greater insulin resistance included waist circumference and heroin use. Factors independently associated with abnormal glucose tolerance (impaired glucose tolerance or diabetes) included age >or=55 years and Hispanic ethnicity. HIV-infected men with diabetes risk factors should undergo screening for diabetes regardless of HAART use. Interventions targeting modifiable risk factors, including overweight and physical inactivity, are warranted. The potential impact of opiate and alcohol abuse on glucose metabolism should be recognized in clinical care, and addressed in future research studies of HIV-infected persons.


A Fringe-modified Notch signal affects specification of mesoderm and endoderm in the sea urchin embryo.
Fringe proteins are O-fucose-specific beta-1,3 N-acetylglucosaminyltransferases that glycosylate the extracellular EGF repeats of Notch and enable Notch to be activated by the ligand Delta. In the sea urchin, signaling between Delta and Notch is known to be necessary for specification of secondary mesenchyme cells (SMCs). The Lytechinus variegatus Fringe homologue is expressed in both the signaling and receiving cells during this first Delta-Notch signal. Perturbation of Fringe expression through morpholino antisense oligonucleotide (MO) injection results in fewer SMCs but also causes decreased and delayed archenteron invagination. Partial endoderm specification occurs but expression of some endoderm genes is compromised. The data are consistent with a Fringe-requiring Notch signal as one upstream component of archenteron morphogenesis. Finally, Fringe perturbations result in more severe phenotypes than those previously reported for Notch dominant-negative (LvN(neg)) injections or reported here for Notch MO (NMO) injections. Injecting a combination of LvN(neg) and NMO results in a more severe phenotype than either treatment alone, and this combination phenocopies the fringe MO embryos. Taken together, the results show that Fringe is necessary both for maternal and zygotic Notch signals, and these Notch signals affect specification of mesoderm and endoderm.


How do Australian podiatrists manage patients with diabetes? The Australian diabetic foot management survey.
Diabetic foot complications are the leading cause of lower extremity amputation and diabetes-related hospitalisation in Australia. Studies demonstrate significant reductions in amputations and hospitalisation when health professionals implement best practice management. Whilst other nations have surveyed health professionals on specific diabetic foot management, to the best of the authors' knowledge this appears not to have occurred in Australia. The primary aim of this study was to examine Australian podiatrists' diabetic foot management compared with best practice recommendations by the Australian National Health Medical Research Council. A 36-item Australian Diabetic Foot Management survey, employing seven-point Likert scales (0 = Never; 7 = Always) to measure multiple aspects of best practice diabetic foot management was developed. The survey was briefly tested for face and content validity. The survey was electronically distributed to Australian podiatrists via professional associations. Demographics including sex, years treating patients with diabetes, employment-sector and patient numbers were also collected. Chi-squared and Mann Whitney U tests were used to test differences between sub-groups. Three hundred and eleven podiatrists responded; 222 (71%) were female, 158 (51%) from the public sector and 11-15 years median experience. Participants reported treating a median of 21-30 diabetes patients each week, including 1-5 with foot ulcers. Overall, participants registered median scores of at least "very often" (>6) in their use of most items covering best practice diabetic foot management. Notable exceptions were: "never" (1 (1 - 3)) using total contact casting, "sometimes" (4 (2 - 5)) performing an ankle brachial index, "sometimes" (4 (1 - 6)) using University of Texas Wound Classification System, and "sometimes" (4 (3 - 6) referring to specialist multi-disciplinary foot teams. Public sector podiatrists reported higher use or access on all those items compared to private sector podiatrists (p < 0.01). This study provides the first baseline information on Australian podiatrists' adherence to best practice diabetic foot guidelines. It appears podiatrists manage large caseloads of people with diabetes and are generally implementing best practice guidelines recommendations with some notable exceptions. Further studies are required to identify barriers to implementing these recommendations to ensure all Australians with diabetes have access to best practice care to prevent amputations.


Experience with modified remotely controlled fluoroscopic equipment for gastrointestinal examination in debilitated patients.
The concept and realization of a highly automated remotely controlled fluoroscopic system, applicable to the examination of debilitated or uncooperative patients, have been described. The addition of a rotating cradle, remotely controlled barium administration, magazine-fed roll film camera, remotely inflated paddle for prone-pressure spot filming, and a vacuum restraining device have been described for the examination of uncooperative as well as cooperative patients. These patients can be examined with as high a degree of sophistication and automation as is now available. Future developments in this area may lie in the realm of multitable remotely controlled fluoroscopic rooms, operation of the above equipment by a super technologist, programmed gastrointestinal examinations utilizing the above equipment and automated programming, and teleremote controlled fluoroscopy from a centralized location with coaxial cable or microwave transmission.


Real-world direct-acting antiviral treatment in kidney transplant and hemodialysis patients: the EpiTer-2 multicenter observational study.
Patients who undergo hemodialysis (HD) or kidney transplantation (KTx) previously had limited possibilities for treatment of hepatitis C virus (HCV) infection. Direct-acting antivirals (DAA) give these patients a chance of virus eradication and safe transplantation. The aim of this study was to evaluate the effectiveness and safety of DAA in KTx and HD patients in real-world settings. Sustained virologic response (SVR) and treatment safety were analyzed in KTx and HD patients from the EpiTer-2 database, which included HCV-infected subjects treated with DAA between 2015 and 2019. Additionally, for KTx patients, changes in creatinine concentration, estimated glomerular filtration rate (eGFR), proteinuria within a year after treatment, and changes in the need for calcineurin inhibitors were assessed. Among 10,152 patients from the EpiTer-2 database 148 were selected, 85 after KTx and 63 undergoing HD. The most common genotype, 1b HCV, was found in 73% and 86% of patients, respectively. Cirrhosis was noted in 10% and 19%, respectively. The most common DAA regimen after KTx was sofosbuvir/ledipasvir (54%), whereas in HD patients it was ombitasvir/paritaprevir/ritonavir +/- dasabuvir (56%). All patients with available follow-up results achieved SVR. No deaths, kidney loss or acute rejection episodes were noted. The most common adverse effects in both groups were anemia and weakness. One year after treatment, creatinine concentration, eGFR and proteinuria remained stable in the majority of patients. DAA treatment of HCV infection demonstrated high effectiveness and safety in hemodialyzed patients and patients who had undergone KTx in this real-world study.


CRISPR Technology for Genome Activation and Repression in Mammalian Cells.
Targeted modulation of transcription is necessary for understanding complex gene networks and has great potential for medical and industrial applications. CRISPR is emerging as a powerful system for targeted genome activation and repression, in addition to its use in genome editing. This protocol describes how to design, construct, and experimentally validate the function of sequence-specific single guide RNAs (sgRNAs) for sequence-specific repression (CRISPRi) or activation (CRISPRa) of transcription in mammalian cells. In this technology, the CRISPR-associated protein Cas9 is catalytically deactivated (dCas9) to provide a general platform for RNA-guided DNA targeting of any locus in the genome. Fusion of dCas9 to effector domains with distinct regulatory functions enables stable and efficient transcriptional repression or activation in mammalian cells. Delivery of multiple sgRNAs further enables activation or repression of multiple genes. By using scaffold RNAs (scRNAs), different effectors can be recruited to different genes for simultaneous activation of some and repression of others. The CRISPRi and CRISPRa methods provide powerful tools for sequence-specific control of gene expression on a genome-wide scale to aid understanding gene functions and for engineering genetic regulatory systems.


B cells capturing antigen conjugated with CpG oligodeoxynucleotides induce Th1 cells by elaborating IL-12.
APCs initiate T cell-mediated immune responses against foreign Ags. Dendritic cells are professional APCs that play unique roles, including Ag-nonspecific capture, priming of naive T cells, and Th1 induction, whereas B cells generally lack these functions. In this study we uncovered novel aspects of murine B cells as APCs using CpG oligodeoxynucleotides (CpG) conjugated with an Ag. B cells served as efficient APCs independently of surface Igs. This characteristic was underlaid by the CpG-mediated Ag uptake and presentation, which were functional only when CpG were covalently conjugated to Ag. The B cells cultured with CpG-conjugated Ag not only enhanced IFN-gamma formation by Th1 cells, but also induced Th1 differentiation from unprimed T cells. These effects paralleled with the increase in the expression of CD40, CD86, and class II molecules on B cells and the coordinated production of IL-12 by the cells. To our knowledge this is the first report revealing that B cells share with dendritic cells common intrinsic characteristics, such as the Ag-nonspecific capture and presentation, and the induction of Th1 differentiation from unprimed T cells.


Deprivation and poor health in rural areas: inequalities hidden by averages.
Poor health and social deprivation scores in 570 wards in East Anglia, UK, were much less associated in rural than in urban areas. The deprivation measure most closely related to poor health in the least accessible rural wards was male unemployment, but use of this measure did not remove the urban-rural gradient of association strength. Neither did replacing wards by smaller enumeration districts as the units of analysis. The differences between urban and rural correlations were removed by restricting the comparison to wards with the same unemployment range and combining pairs of rural wards with similar deprivation values. Apparent differences between rural and urban associations are therefore not due to the choice of deprivation indices or census areas but are artifacts of the greater internal variability, smaller average deprivation range and smaller population size of rural small areas. Deprived people with poor health in rural areas are hidden by favourable averages of health and deprivation measures and do not benefit from resource allocations based on area values.


Paraquat toxicity and oxidative damage. Reduction by melatonin.
The ability of melatonin to protect against paraquat-induced oxidative damage in rat lung, liver, and serum was examined. Changes in the levels of malondialdehyde (MDA) plus 4-hydroxyalkenals (4-HDA) and reduced and oxidized glutathione concentrations were measured. Paraquat (50 mg/kg) was injected i.p. into either Sprague-Dawley or Wistar rats with or without the co-administration of 5 mg/kg melatonin. Paraquat alone increased MDA + 4-HDA levels in serum and lungs of both rat strains, with these increases being abolished by melatonin co-treatment. Paraquat also decreased reduced glutathione levels and increased oxidized glutathione concentrations in lung and liver; these changes were negated by melatonin. The effect of melatonin on paraquat-induced mortality was also studied. Paraquat at a dose of 79 mg/kg was lethal for 50% of animals within 24 hr; when administered together with melatonin, the LD50 for paraquat increased to 251 mg/kg.


Equilibrative nucleoside transporter 1 gene polymorphisms and clinical outcomes following acute coronary syndromes: findings from the PLATelet inhibition and patient Outcomes (PLATO) study.
In the PLATelet inhibition and patient Outcomes (PLATO) study, the P2Y12 inhibitors ticagrelor and clopidogrel were compared in the treatment of acute coronary syndromes (ACS). Ticagrelor was shown to reduce occurrence of the primary end point - a composite of death from vascular causes, myocardial infarction, or stroke - compared to clopidogrel. Ticagrelor's pleiotropic effects on reuptake of adenosine via inhibition of equilibrative nucleoside transporter 1 (ENT1) have been hypothesized to contribute to this. Several polymorphisms of ENT1 are known to exist. We explored the interaction between ENT1 polymorphisms and clinical outcomes in ACS patients participating in the PLATO genetic substudy. Using genotyping data obtained in a genome-wide association study, the gene region encoding ENT1 was assessed and 94 polymorphisms were identified. After quality control filtering, data from 9943 participants were included. Subjects were divided into discovery (phase 1, n = 3970) and replication (phase 2, n = 5973) cohorts. Cox-regression analysis of the relationship between variants and seven efficacy and safety outcomes was performed in discovery, replication, and combined cohorts. Treatment-marker interactions were also determined. Although 35 variants were found with associations to the investigated outcomes reaching p < 0.05 in the discovery cohort, only one of these was replicated in phase 2 of the analysis and also reached the predetermined level of statistical significance in the combined data, taking into account the number of tests performed: the rare polymorphism rs141034817, with a frequency of 0.2%, was significantly associated with bleeding. Thirty-three treatment-marker interactions were found with a significance level of p < 0.05 in phase 1, but none was replicated in phase 2. We found no significant interaction between ENT1 genotype and clinical outcomes in ACS patients treated with ticagrelor or clopidogrel, apart from the association between a rare polymorphism and bleeding that requires further study. If ticagrelor's pleiotropic effects on adenosine uptake are clinically relevant, these do not appear to be significantly affected by variation in the ENT1 gene.


Phosphate analogues in the dissection of mechanism.
Phosphoryl group transfer is central to genetic replication, cellular signalling and many metabolic processes. Understanding the mechanisms of phosphorylation and phosphate ester and anhydride cleavage is key to efforts towards biotechnological and biomedical exploitation of phosphate-handling enzymes. Analogues of phosphate esters and anhydrides are indispensable tools, alongside protein mutagenesis and computational methods, for the dissection of phosphoryl transfer mechanisms. Hydrolysable and non-hydrolysable phosphate analogues have provided insight into the nature and sites of phosphoryl transfer processes. Kinetic isotope effects and crystallography using transition state analogues have painted more detailed pictures of transition states and how enzymes work to stabilise them.


The validity of the mortality to incidence ratio as a proxy for site-specific cancer survival.
The complement of the cancer mortality to incidence ratio [1-(M/I)] has been suggested as a valid proxy for 5-year relative survival. Whether this suggestion holds true for all types of cancer has not yet been adequately evaluated. We used publicly available databases of cancer incidence, cancer mortality and relative survival to correlate relative survival estimates and 1-(M/I) estimates from Denmark, Finland, Iceland, Norway, Sweden, the USA and the Netherlands. We visually examined for which tumour sites 5-year relative survival cannot simply be predicted by the 1-(M/I) and evaluated similarities between countries. Country-specific linear regression analyses show that there is no systematic bias in predicting 5-year relative survival by 1-(M/I) in five countries. There is a small but significant systematic underestimation of survival from prognostically poor tumour sites in two countries. Furthermore, the 1-(M/I) overestimates survival from oral cavity and liver cancer with >10% in at least two of the seven countries. By contrast, the proxy underestimates survival from soft tissue, bone, breast, prostate and oesophageal cancer, multiple myeloma and leukaemia with >10% in at least two of the seven countries. The 1-(M/I) is a good approximation of the 5-year relative survival for most but not all tumour sites.


The innervation of the middle ear muscles of the rat.
The innervation of the tensor tympani muscle and the stapedius muscle in the rat was studied. This was done by acetylcholinesterase in toto staining of the tympanic bullae and of muscles dissected separately, acetylcholinesterase staining of serial cross-sections of the muscles, silver impregnation of serial sections of complete tympanic bullae, serial semithin sections stained according to Laczko & Levai and electron microscopy of both muscles. The gross innervation of the muscles and the relation to other nerves in the bulla are described. It is shown that both muscles are innervated by very thin nerve fibres which form a well-organised elaborate network in the muscles, with very short branches that connect with motor endplates. Electron microscopically there are indications that the endplates in the stapedius muscle seem to enable faster activation of the muscle fibres than those of tensor tympani muscle. No morphological evidence for any sensory innervation of the muscles could be detected in the muscles themselves, in the connective tissue related to the muscles, or in the contents of the bulla tympanica. It is postulated that the afferent input of the acoustic middle ear muscle reflex is sound alone and that sensory information from the muscles themselves or from other structures in the tympanic bulla do not contribute to the reflex.


Correction.
It has been called to our attention that our announcement of the granting of the Third Intermediate Sugar Research Foundation Award to Leslie F. Wiggins (Science, May 14, p. 502) was incorrect in several respects. The Imperial College of Tropical Agriculture, mentioned in the item as being located in Kingston, Jamaica, and as having been "recently created," is actually located in Trinidad, B.W.I., and has been in existence for 27 years. We learn that it was founded as the West Indian Agricultural College in 1921 and that its name was changed to its present one in 1924. It was also stated in the announcement that Dr. Wiggins had been appointed research director at the College. His appointment was as director of sugar research under a new Sugar Research Scheme launched in 1947. We regret that these misstatements occurred.


[Cave: interscalene catheters].
Interscalene regional anesthesia is an established and highly effective procedure; however, it represents an increased level of risk due to the close proximity of anatomical structures, such as the cervical spinal cord and many vessels. Furthermore, due to inadvertent placement of a catheter close to the cervical spinal cord or into a vessel, as opposed to a single shot injection technique, it remains a latent danger until it is removed. This article describes seven cases of misplaced catheters. The etiology and symptoms are discussed as well as recommendations regarding the prevention of catastrophic complications. As a result, internal practice guidelines are recommended for anesthesia departments in order to enhance the safety and quality of regional anesthesia.


Tolerability of high doses of lercanidipine versus high doses of other dihydropyridines in daily clinical practice: the TOLERANCE Study.
The TOLERANCE study was aimed to compare the tolerability of high doses of lercanidipine (20 mg) with that of other frequently used dihydropyridines (amlodipine 10 mg/nifedipine GITS 60 mg) in the treatment of essential hypertension in daily clinical practice. It was an observational, transversal, multicentre study performed in a Primary Care Setting. A total of 650 evaluable patients with essential hypertension and age > or = 18 years were included. They had been treated with high doses of lercanidipine (n= 446) or amlodipine/nifedipine GITS (n= 204) during at least 1 month and previously with low doses (10 mg, 5 mg, and 30 mg, respectively) of the same drugs. The main objective was to compare the rates of vasodilation-related adverse events between both groups. Rates of signs and symptoms related to vasodilation were significantly higher (P < 0.001) in the amlodipine/nifedipine GITS group (76.8%, CI 95%[70.7; 82.9]) than in lercanidipine group (60.8%, [56.1;65.5]). Blood pressure control (< 140/90 mmHg or <130/80 for diabetics) and type of concomitant antihypertensive medications were similar in both groups. Treatment compliance was good (around 93%) and fairly comparable in both groups. Most adverse events with lercanidipine were mild (74.5% vs. 64% in amlodipine/nifedipine GITS group, P= 0.035) whereas severe adverse event rates did not differ significantly between groups (2.8% vs. 3.6%). In conclusion, treatment with lercanidipine at high doses is associated with a lower rate of adverse events related to vasodilation compared to high doses of amlodipine or nifedipine GITS in clinical practice.


Postal surveys: a review of articles published in the Medical Journal of Malaysia between 1991-2000.
Postal surveys are becoming an increasingly popular method of conducting research in Malaysia. However, the response rates from these surveys have not been well studied. All postal surveys published in the Medical Journal of Malaysia between 1991-2000 were therefore reviewed to provide an overview of the response rates that may be expected, and to assess if this is a valid method of conducting research here. The response rates of postal surveys ranged from 30-87.5%. Seven of 13 postal surveys (54%) had a response rate of less than 50% and only 2 of 13 (15.4%) had a response rate of more than 70%, raising doubts on the use of postal surveys as a research tool.


Nicotine concentration in leaves of flue-cured tobacco plants as affected by removal of the shoot apex and lateral buds.
It is believed that the nicotine concentration in tobacco is closely correlated with the amount of nitrogen (N) supplied. On the other hand, N uptake mainly occurs at the early growth stage, whereas nicotine concentration increases at the late growth stage, especially after removing the shoot apex. To identify the causes of the increased nicotine concentration in tobacco plants, and to compare the effects of different ways of mechanical wounding on nicotine concentration, field experiments were carried out in Fuzhou, Fujian Province in 2003 and 2004. Excision of the shoot apex had almost no influence on N content in the plant; however, it caused dramatic increases in nicotine concentration in leaves, especially in the middle and upper leaves. An additional increase of the nicotine concentration was obtained by removal of axillary buds. The wounding caused by routine leaf harvests, however, did not change the leaf nicotine concentration, and neither did reducing leaf harvest times. The present results revealed no direct relationship between N supply and nicotine concentration in tobacco leaves, and indicate that not all kinds of mechanical wounding were capable of stimulating nicotine synthesis in tobacco plants. Since nicotine production is highly dependent on the removal of apical meristems and hence on the major sources of auxin in the plant, and application of 1-naphthylacetic acid onto the cut surface of the stem after removing the shoot apex markedly decreased the nicotine concentration in different leaves and the total nicotine content in the plant, the results suggest that decreased auxin supply caused by removal of the shoot apex as a kind of mechanical wounding might regulate nicotine synthesis in the roots of tobacco plants.


Solid Waste Management Policy Implications on Waste Process Choices and Systemwide Cost and Greenhouse Gas Performance.
Solid waste management (SWM) is a key function of local government and is critical to protecting human health and the environment. Development of effective SWM strategies should consider comprehensive SWM process choices and policy implications on system-level cost and environmental performance. This analysis evaluated cost and select environmental implications of SWM policies for Wake County, North Carolina using a life-cycle approach. A county-specific data set and scenarios were developed to evaluate alternatives for residential municipal SWM, which included combinations of a mixed waste material recovery facility (MRF), anaerobic digestion, and waste-to-energy combustion in addition to existing SWM infrastructure (composting, landfilling, single stream recycling). Multiple landfill diversion and budget levels were considered for each scenario. At maximum diversion, the greenhouse gas (GHG) mitigation costs ranged from 30 to 900 $/MTCO2e; the lower values were when a mixed waste MRF was used, and the higher values when anaerobic digestion was used. Utilization of the mixed waste MRF was sensitive to the efficiency of material separation and operating cost. Maintaining the current separate collection scheme limited the potential for cost and GHG reductions. Municipalities seeking to cost-effectively increase landfill diversion while reducing GHGs should consider waste-to-energy, mixed waste separation, and changes to collection.


Live in vivo imaging of Egr-1 promoter activity during neonatal development, liver regeneration and wound healing.
The zinc finger transcription factor Egr-1 (Early growth response 1) is central to several growth factors and represents an important activator of target genes not only involved in physiological processes like embryogenesis and neonatal development, but also in a variety of pathophysiological processes, for example atherosclerosis or cancer. Current options to investigate its transcription and activation in vivo are end-point measurements that do not provide insights into dynamic changes in the living organism. We developed a transgenic mouse (Egr-1-luc) in which the luciferase reporter gene is under the control of the murine Egr-1 promoter providing a versatile tool to study the time course of Egr-1 activation in vivo. In neonatal mice, bioluminescence imaging revealed a high Egr-1 promoter activity reaching basal levels three weeks after birth with activity at snout, ears and paws. Using a model of partial hepatectomy we could show that Egr-1 promoter activity and Egr-1 mRNA levels were increased in the regenerating liver. In a model of wound healing, we demonstrated that Egr-1 promoter activity was upregulated at the site of injury. Taken together, we have developed a transgenic mouse model that allows real time in vivo imaging of the Egr-1 promoter activity. The ability to monitor and quantify Egr-1 activity in the living organism may facilitate a better understanding of Egr-1 function in vivo.


Subcellular localization of proline-rich tyrosine kinase 2 during oocyte fertilization and early-embryo development in mice.
Proline-rich tyrosine kinase 2 (Pyk2), a non-receptor tyrosine kinase, is a member of the focal adhesion kinase family and is highly expressed in oocytes. Using a combination of confocal microscopy and RNAi, we localized and studied the function of both Pyk2 and tyrosine-phosphorylated Pyk2 (p-Pyk2) during mouse oocyte fertilization and early embryo development. At the onset of fertilization, Pyk2 and p-Pyk2 were detected predominantly in sperm heads and the oocyte cytoplasm. Upon formation of male and female pronuclei, Pyk2 and its activated form leave the cytoplasm and accumulate in the two pronuclei. We detected Pyk2 in blastomere nuclei and found both Pyk2 and p-Pyk2 in the pre-blastula cytoplasm. Pyk2 and its activated form then disappeared from the blastula nuclei and localized to the perinuclear regions, where blastula cells come into contact with each other. Pyk2 knockdown via microinjection of siRNA into the zygote did not inhibit early embryo development. Our results suggest that Pyk2 plays multiple functional roles in mouse oocyte fertilization as well as throughout early embryo development.


Regulation of NAD(P)H:quinone oxidoreductase 1 gene expression by CYP1A1 activity.
The dioxin 2,3,7,8-tetrachlorodibenzo-para-dioxin (TCDD) induces phase I and II xenobiotic metabolizing enzymes (XME) which act sequentially to eliminate different classes of xenobiotics. The transcriptional effects of TCDD are generally mediated by the arylhydrocarbon receptor (AhR). We hypothesized that TCDD could also act indirectly, by increasing the activity of cytochrome P450 1A1 (CYP1A1), a phase I gene, which could then mediate the induction of other XME genes, such as the NAD(P)H:quinone oxidoreductase 1 (NQO1). To test this hypothesis, NQO1 gene expression was monitored after either overexpression of CYP1A1 or siRNA-mediated knock-down of CYP1A1 activity in the hepatoma cell line HepG2. Overexpression of CYP1A1 in the absence of TCDD was carried out using either adenoviral infection or the "Tet-off" system. Recombinant adenoviruses were produced encoding no protein, CYP1A1 (Ad1A1), or a mutated inactive CYP1A1 (Ad1A1mut). In the HepG2 Tet-off cell line, CYP1A1 expression was induced by the removal of doxycycline (dox) from the cell medium. Ad1A1 infection or dox removal induced CYP1A1 activity and H(2)O(2) production similarly to TCDD treatment. Moreover, in both systems, the amount of NQO1 mRNA increased to the same level as after TCDD treatment (approximately 2-fold). The UDP-glucuronosyl transferase 1A6 (UGT1A6) gene is also similarly regulated. NQO1 gene expression was not induced when mutant, inactive CYP1A1 was overexpressed or when the antioxidant N-acetyl cysteine (NAC) was added to Ad1A1. Finally, either NAC or siRNA directed against CYP1A1 mRNA decreased the induction of NQO1 gene expression by TCDD. We conclude that, after exposure to TCDD, the NQO1 gene expression can be controlled by CYP1A1 activity through an oxidative stress mediated pathway.


Lipidomics profiling reveals distinct patterns of plasma sphingolipid alterations in Alzheimer's disease and vascular dementia.
Alzheimer's disease (AD) and vascular dementia (VaD) are two of the commonest causes of dementia in the elderly. Of the myriad biomolecules implicated in dementia pathogenesis, sphingolipids have attracted relatively scant research attention despite their known involvement in multiple pathophysiological processes. The potential utility of peripheral sphingolipids as biomarkers in dementia cohorts with high concomitance of cerebrovascular diseases is also unclear. Using a lipidomics platform, we performed a case-control study of plasma sphingolipids in a prospectively assessed cohort of 526 participants (non-cognitively impaired, NCI = 93, cognitively impaired = 217, AD = 166, VaD = 50) using a lipidomics platform. Distinct patterns of sphingolipid alterations were found in AD and VaD, namely an upregulation of d18:1 species in AD compared to downregulation of d16:1 species in VaD. In particular, GM3 d18:1/16:0 and GM3 d18:1/24:1 showed the strongest positive associations with AD. Furthermore, evaluation of sphingolipids panels showed specific combinations with higher sensitivity and specificity for classification of AD (Cer d16:1/24:0. Cer d18:1/16:0, GM3 d16:1/22:0, GM3 d18:1/16:0, SM d16:1/22:0, HexCer d18:1/18:0) and VAD (Cer d16:1/24:0, Cer d18:1/16:0, Hex2Cer d16:1/16:0, HexCer d18:1/18:0, SM d16:1/16:0, SM d16:1/20:0, SM d18:2/22:0) compared to NCI. AD and VaD are associated with distinct changes of plasma sphingolipids, warranting further studies into underlying pathophysiological mechanisms and assessments of their potential utility as dementia biomarkers and therapeutic targets.


The Multivariate Physical Activity Signatures Associated With Self-Regulation, Executive Function, and Early Academic Learning in 3-5-Year-Old Children.
The evidence regarding associations between intensity-specific physical activity and cognitive and learning outcomes in preschoolers is inconsistent and limited by low sample sizes and analytical approaches that cannot handle the multicollinearity among multiple physical activity intensity variables. We aimed to determine the multivariate physical activity intensity signatures associated with self-regulation, executive function, and early academic learning in preschool children aged 3-5 years. A 711 Norwegian preschool children (mean age 4.6 years, 52% boys) provided valid data on physical activity (ActiGraph GT3X+), self-regulation, executive function, and early academic learning during 2019-2020. Multivariate pattern analysis was used to determine associations between uniaxial and triaxial intensity spectra (time spent in intensities from 0-99 to ≥15,000 counts per minute) and the outcomes in the total sample and in subgroups split by sex and age (median split). Uniaxial data led to the highest explained variances (R 2) and were reported as the primary findings. We found significant association patterns between physical activity and numeracy (R 2 = 4.28%) and inhibition (R 2 = 1.48%) in the total sample. The associations with numeracy were negative for time spent sedentary (0-99 counts per minute) and positive for time spent in moderate to vigorous intensities (≥ 1,000 counts per minute). The associations with inhibition were positive for time spent sedentary (0-99 counts per minute) and in vigorous intensities (≥ 8,500 counts per minute) and negative for time spent in low to moderate intensities (100-3,499 counts per minute). Associations with numeracy were stronger in boys (R 2 = 5.58%) and older children (R 2 = 7.27%), and associations with inhibition were stronger in girls (R 2 = 3.12%) and older children (R 2 = 3.33%). In conclusion, we found weak associations with numeracy and inhibition across the physical activity intensity spectrum in preschool children.


Increased blood viscosity in diabetic proliferative retinopathy.
Blood rheology and haemostasis were assessed in 18 diabetics with proliferative retinopathy and in 18 diabetics without proliferative retinopathy, matched for age, sex, smoking habit and type, duration and treatment of diabetes. Proliferative retinopathy was associated with significantly higher levels of blood viscosity at high and low shear rates, which were related to higher levels of plasma viscosity and fibrinogen. Blood urea, glucose, glycosylated haemoglobin and white cell count were also significantly higher, whereas haematocrit, red cell deformability and several other haematological and biochemical variables did not differ significantly in the 2 groups. In view of these findings, and of our recent demonstration that increased blood viscosity also exists in those patients with retinal vein occlusion who develop a similar proliferative retinopathy, we suggest that hyperviscosity may contribute to retinal ischaemia and hence proliferative retinopathy.


Genesis of 1,2-dimethylhydrazine-induced colon cancer. A light and electron microscopic study.
Rats treated with 1,2-dimethylhydrazine (DMH) exhibited colonic mucosal dysplastic foci prior to the development of tumors. Ultrastructurally, these, as well as the cancers that subsequently developed, were characterized by alterations in plasma membranes and an increase in cytoplasmic ribosomal particles, principally in stem cells and their absorptive derivatives. Rare Kulchitsky cells appeared preserved, but the mucin-producing or goblet-cell elements were compressed and atrophic. In addition, nuclear aberrations were more pronounced in the cancer than in the dysplastic foci. The principal ultrastructural difference between the so-called well-differentiated and mucinous forms of DMH-induced cancers was the presence of frequent intracytoplasmic lumens in the mucinous form. Such structures represented the analogues of signet ring cells observed by light microscopy. This experimental model of human colonic cancer shows that the mucosal stem-cell and dysplastic lesions represent their cytogenetic and histogenetic progenitors.


Effect of instillation of eyedrops for dry eye on optical quality.
To investigate the effects of viscosity and suspensibility of eyedrops for dry eye by evaluating an eyedrop with one of the solutions or no solution (0.3% sodium hyaluronate ophthalmic solution, 3% diquafosol ophthalmic solution, and 2% rebamipide ophthalmic suspension) on ocular higher-order aberrations (HOAs) and forward light scatter. We evaluated ocular HOAs and forward light scatter before and 1, 5, and 10 minutes after instillation of three eyedrops for dry eye in 15 healthy subjects. Saline served as the control. The HOAs were measured for a 4-mm pupil using a wavefront sensor. The obtained aberration data were analyzed in the central 4-mm diameter for total HOAs up to the sixth-order Zernike polynomials. Forward light scatter was quantified with a straylight meter. A significant increase was seen in the HOAs 1 minute after instillation of the three eyedrops for dry eye; the HOAs recovered to the baseline level thereafter. When 0.3% sodium hyaluronate was compared with 2% rebamipide and 3% diquafosol, the HOAs increased significantly (P < 0.01 for both comparisons) immediately after instillation. A significant increase in forward light scatter occurred 1 minute after instillation of rebamipide suspension and returned to the preinstillation level 5 minutes after instillation. No significant changes in forward light scatter occurred after instillation of 3% diquafosol or 0.3% sodium hyaluronate. Quantitative serial measurement of HOAs and forward light scatter showed that the temporal reduction in optical quality may be attributed mainly to increased HOAs after instillation of highly viscous 0.3% sodium hyaluronate ophthalmic solution and to increased forward light scatter after instillation of 2% rebamipide ophthalmic suspension in healthy subjects.


RAD-ical New Insights into RAD51 Regulation.
The accurate repair of DNA is critical for genome stability and cancer prevention. DNA double-strand breaks are one of the most toxic lesions; however, they can be repaired using homologous recombination. Homologous recombination is a high-fidelity DNA repair pathway that uses a homologous template for repair. One central HR step is RAD51 nucleoprotein filament formation on the single-stranded DNA ends, which is a step required for the homology search and strand invasion steps of HR. RAD51 filament formation is tightly controlled by many positive and negative regulators, which are collectively termed the RAD51 mediators. The RAD51 mediators function to nucleate, elongate, stabilize, and disassemble RAD51 during repair. In model organisms, RAD51 paralogs are RAD51 mediator proteins that structurally resemble RAD51 and promote its HR activity. New functions for the RAD51 paralogs during replication and in RAD51 filament flexibility have recently been uncovered. Mutations in the human RAD51 paralogs (RAD51B, RAD51C, RAD51D, XRCC2, XRCC3, and SWSAP1) are found in a subset of breast and ovarian cancers. Despite their discovery three decades ago, few advances have been made in understanding the function of the human RAD51 paralogs. Here, we discuss the current perspective on the in vivo and in vitro function of the RAD51 paralogs, and their relationship with cancer in vertebrate models.


[Clinical manifestations of APS-nephropathy in primary antiphospholipid syndrome].
To elicit clinical features of nephropathy associated with antiphospholipid syndrome (APSN) in patients with primary antiphospholipid syndrome (PAPS). The analysis of clinical characteristics and course of APSN has covered 24 patients with PAPS (16 females and 8 males, mean age 34.3 years). Renal damage was represented by arterial hypertension (AH), urinary syndrome, functional decline. All the patients were tested for anticardiolipin antibodies and/or lupus anticoagulant. Renal biopsy was made in 7 patients. PAPS patients developed renal affection in the onset of APS or within the first 5 years of its course. In the majority of patients APSN combined with abnormalities of CNS, heart and skin. Arterial/arteriolar thromboses prevailed. APSN manifested with: AH (n = 23, severe AH in 11), abnormal renal filtration (n = 17, creatinine rise in 8), urinary syndrome with proteinuria (n = 23, in 14 with hematuria). The following clinical variants of APSN were proposed: urinary syndrome with AH (n = 16; 67%), acute nephritic syndrome (n = 7; 29%), nephrotic syndrome (n = 1). Morphological studies of biopsies from APSN patients have revealed sclerotic changes, thrombotic microangiopathy, nonspecific alterations in the glomeruli. APSN is a variant of microvascular renal affection caused by thrombotic processes in intra-organ microcirculation. It is an early clinical marker of APS. Clinically, APSN manifests with vascular renal affection, the earliest symptom being inhibition of glomerular filtration. Clinical combinations of the symptoms allow to distinguish variant of APSN suggesting the existence of acute and chronic APSN. Combination of APSN with affection of the CNS, heart and skin points to a special PAPS subtype characterized by generalized ischemic damage to the organs as a result of intraorganic arterial and/or arteriolar thromboses.


Development and optimization of a gas chromatography/mass spectrometry method for the analysis of thermochemolytic degradation products of phthiocerol dimycocerosate waxes found in Mycobacterium tuberculosis.
The phthiocerol dimycocerosates (PDIMs) are certain stable and hydrophobic waxes found in the cell membrane of Mycobacterium tuberculosis, bacteria that cause an infectious disease of growing concern worldwide. Previous studies report the analysis of derivatives of the hydrolysed PDIMs from biological samples, following complex extraction and offline derivatization of PDIMs biomarkers, prior to their analysis by gas chromatography/mass spectrometry (GC/MS). We developed and optimized a GC/MS method based on selected ion monitoring (SIM) to detect the derivatives produced via the thermally assisted hydrolysis and methylation (THM) of the PDIMs from the cell membrane of M. tuberculosis. The extraction of PDIMs from culture is simple, and their thermochemolysis is carried out automatically online, thus avoiding the time-consuming derivatization steps of hydrolysis and esterification, usually performed offline. For standard PDIMs in petroleum ether, our optimized method gave an excellent linearity (R(2) = 0.99) at concentrations between 0.172 and 27.5 ng/mL, a good precision (RSD = 11.42%), and a limit of detection (LOD) of 100 pg/mL. For the PDIMs extracted from dilutions of M. tuberculosis culture, the method gave good linearity (R(2) = 0.9685) and an estimated LOD of 400 CFU/mL (CFU = colony forming units) in sterile distilled water. A GC/MS(SIM) method is presented for the rapid and quantitative detection of M. tuberculosis, based on the online thermochemolysis of lipidic biomarkers extracted from the bacterial culture. The method has the potential to be applied in human and veterinary clinical laboratories for the rapid diagnosis of tuberculosis in infected biological samples.


A case-control study on the relationship between salt intake and salty taste and risk of gastric cancer.
To investigate the relationship between salt intake and salty taste and risk of gastric cancer. A 1:2 matched hospital based case-control study including 300 patients with gastric cancer and 600 cancer-free subjects as controls. Subjects were interviewed with a structured questionnaire containing 80 items, which elicited information on dietary, lifestyle habits, smoking and drinking histories. Subjects were tested for salt taste sensitivity threshold (STST) using concentrated saline solutions (0.22-58.4 g/L). Conditional logistic regression was used to calculate odds ratios (ORs) and 95% confidence intervals (95% CI). Alcohol and tobacco consumption increased the risk of gastric cancer [OR (95% CI) was 2.27 (1.27-4.04) for alcohol and 2.41 (1.51-3.87) for tobacco]. A protective effect was observed in frequent consumption of fresh vegetable and fruit [OR (95% CI) was 0.92 (0.58-0.98) for fresh vegetable and 0.87 (0.67-0.93) for fruit]. Strong association was found between STST ≥ 5 and gastric cancer [OR = 5.71 (3.18-6.72)]. Increased STST score was significantly associated with salted food intake and salty taste preference (P < 0.05). A high STST score is strongly associated with gastric cancer risk. STST can be used to evaluate an inherited characteristic of salt preference, and it is a simple index to verify the salt intake in clinic.


A rural-urban comparison of prehospital emergency medical services in Nebraska.
Provision of emergency medical services (EMS) in the rural United States presents a unique challenge. While rural and urban EMS outcomes have been compared, differing urban-rural population characteristics and roles for rural ambulance teams can confound such comparisons. A year-long study of the prehospital EMS was conducted in rural Richardson County, Nebraska. Data were collected on the age, sex, and race of patients, response time, transport distance, medical problems encountered, and treatment rendered enroute. These data compared with those from an urban Lancaster County comparison group and statistical data from the Nebraska State Health Department. In the rural county, 70 percent of calls involved the elderly, whereas 38 percent of the urban calls and 36 percent of the Nebraska State calls involved the elderly. The rural ambulance service was more likely to provide for routine transfers, to involve patients with fractures and cardiorespiratory and neurologic problems, and twice as likely to result in hospital admission than was the urban ambulance service. The frequency with which advanced life support measures were applied in the rural area was similar to that in the urban area. The rural area response times were equivalent to the urban area response times after the rural area long-distance transfers were excluded. The location of service in the rural area was more likely to be the hospital or nursing home, whereas the urban location was more likely to be a home, on a highway, or in a public setting. Prehospital EMS in this rural location involved a predominantly elderly population with a large number of routine transfers linking the nursing home and community hospital. Further comparisons of rural and urban EMS outcomes should account for possible differences in type and severity of illness and type and location of service.