Screening the oxidative potential of several mono- and di-halogenated biphenyls and biphenyl ethers in rat hepatocytes.
The present study was designed to investigate the potential of reactive oxygen species (ROS) generating and subsequent ROS-mediated lipid peroxidation (LPO) inducing effect of several mono- and di-halogenated biphenyls and biphenyl ethers in rat hepatocytes in vitro. For this aim, 4-chloro- and 4-bromo biphenyl (4-CB and 4-BB), 4-OH, 4'-BB, 4-bromo diphenylether (4-BDE), 4,4'-dichlorobiphenyl (4,4'-DCB), 4,4'-dibromobiphenyl (4,4'-DBB), and 3,4-dichlorobiphenyl (3,4-DCB) were incubated with freshly isolated rat hepatocytes. Their oxidative potential was evaluated by detecting the intracellular ROS formation by oxidant-sensing fluorescent probes (2',7'-dichlorofluorescein diacetate and C(11)-BODIPY(581/591)) using a multiplate reader and determining the levels of eight LPO products (formaldehyde, malondialdehyde, propanal, butanal, pentanal, hexanal, octanal, and nonanal) by a gas chromatography-electron capture detection. 4-BDE was found to be active both in cytoplasm and in the cell membrane in terms of inducing the formation of ROS. Another important finding was the increase in ROS-inducing potential of 4-BB when the same concentration of the hydroxylated derivative, 4-OH,4'-BB, was incubated with hepatocytes. 4-BDE was also found to be the most effective among all tested compounds in inducing LPO where 4-OH, 4'-BB was again more potent than its unmetabolized form, 4-BB. Lactate dehydrogenase leakage analyses indicated that all tested compounds are cytotoxic; 4-BDE caused the highest LDH leakage compared to other mono-halogenated biphenyls tested. Our results suggest that ROS formation by chlorinated biphenyls and mono-hydroxylated bromobiphenyls, and concomitant induction of LPO might be involved in the cytotoxic effects of these industrial pollutants. Similar effects of mono-BDE are also reported, which is a novel observation.


Estimating prion adsorption capacity of soil by BioAssay of Subtracted Infectivity from Complex Solutions (BASICS).
Prions, the infectious agent of scrapie, chronic wasting disease and other transmissible spongiform encephalopathies, are misfolded proteins that are highly stable and resistant to degradation. Prions are known to associate with clay and other soil components, enhancing their persistence and surprisingly, transmissibility. Currently, few detection and quantification methods exist for prions in soil, hindering an understanding of prion persistence and infectivity in the environment. Variability in apparent infectious titers of prions when bound to soil has complicated attempts to quantify the binding capacity of soil for prion infectivity. Here, we quantify the prion adsorption capacity of whole, sandy loam soil (SLS) typically found in CWD endemic areas in Colorado; and purified montmorillonite clay (Mte), previously shown to bind prions, by BioAssay of Subtracted Infectivity in Complex Solutions (BASICS). We incubated prion positive 10% brain homogenate from terminally sick mice infected with the Rocky Mountain Lab strain of mouse-adapted prions (RML) with 10% SLS or Mte. After 24 hours samples were centrifuged five minutes at 200 × g and soil-free supernatant was intracerebrally inoculated into prion susceptible indicator mice. We used the number of days post inoculation to clinical disease to calculate the infectious titer remaining in the supernatant, which we subtracted from the starting titer to determine the infectious prion binding capacity of SLS and Mte. BASICS indicated SLS bound and removed ≥ 95% of infectivity. Mte bound and removed lethal doses (99.98%) of prions from inocula, effectively preventing disease in the mice. Our data reveal significant prion-binding capacity of soil and the utility of BASICS to estimate prion loads and investigate persistence and decomposition in the environment. Additionally, since Mte successfully rescued the mice from prion disease, Mte might be used for remediation and decontamination protocols.


Potentially lethal complications of central venous catheter placement.
Placement of central venous catheters, although often considered to be a relatively safe and "junior"-level procedure, may be associated with life-threatening complications. A recent surgical death associated with placement of a central venous catheter at this Institution led to submission of a questionnaire to pediatric surgeons referenced through the American Pediatric Surgical Association directory regarding knowledge of similar incidents and information regarding catheter placement-related complications. Results to this response, although anecdotal, provided data regarding complications of an acute nature, which fell into the categories of pneumothorax, hydrothorax, cardiac tamponade, and hemothorax. Of 10 children with cardiac tamponade, 7 were infants, and most complications were associated with needle stick for access, with symptoms developing within minutes up to 12 hours after the procedure. Drainage of the tamponade was performed by aspiration alone in 3 cases; surgical drainage in 6 children resulted in survival in 9 of the 10 patients. Hemothorax was described in 19 patients and appeared to be more common in children in the 1- to 6-year age group, usually associated with percutaneous access techniques. Thoracotomy for hemothorax was performed in 16 children with 11 survivors. Vascular injury to subclavian artery, vein, or superior vena caval were noted in most at operation. Although data included in this review are entirely anecdotal and not subject to scientific scrutiny or analysis, certain conclusions appear evident. Inherent risks of central venous catheters are intrinsic and should be discussed with the family in obtaining preoperative consent, including life-threatening risks that may necessitate urgent surgical intervention (by thoracotomy or other means). Certain technical aspects of the procedure should be rigidly followed with an experienced surgeon in attendance throughout the procedure. Rapid evaluation should be performed for any unexplained problems that occur in the operating theatre or during the early postoperative period.


Assessing the remedy: the case for contracts in clinical trials.
Current orthodoxy in research ethics assumes that subjects of clinical trials reserve rights to withdraw at any time and without giving any reason. This view sees the right to withdraw as a simple extension of the right to refuse to participate all together. In this paper, however, I suggest that subjects should assume some responsibilities for the internal validity of the trial at consent and that these responsibilities should be captured by contract. This would allow the researcher to impose a penalty on the subject if he were to withdraw without good reason and on a whim. This proposal still leaves open the possibility of withdrawing without penalty when it is in the subject's best interests to do so. Giving researchers recourse to legal remedy may now be necessary to protect the science, as existing methods used to increase retention are inadequate for one reason or another.


[New protective effect of simvastatin in rats with experimental steatohepatitis].
The intragastric introduction of carbon tetrachloride (CCl4) (0.2 ml/kg in 50% oil suspension, twice a week) and ethyl alcohol (5% solution ad libitum as the only available drink) in rats over a period of four weeks results in the development of inflammation, fibrosis, and fatty dystrophy in the liver. Such a fast formation of liver damage is obviously caused by potentiating effect of alcohol in combination with CCl4. Simultaneous injection of simvastatin (1 mg/kg, intragastrically) in rats with ethanol--CCl4 hepatitis decreased fatty dystrophy and produced certain anticytolytic and anticholestatic effects without potentiation of microsomal oxidation system damage by hepatotoxins. In addition, simvastatin shows hypolipidemic activity, which is manifested primarily by a decrease in the general holesterol level in the blood serum.


Real-world characteristics of elderly patients with overactive bladder in the United States.
Although much has been published about the demographic and clinical characteristics of elderly patients with overactive bladder (OAB) who were enrolled in clinical trials, very little is known about the general population of elderly Americans with OAB. We update this gap in the literature by using real-world data to describe this population. We used Medicare claims and the Medicare Current Beneficiary Surveys from 2006 to 2011 to identify patients with OAB. We describe the demographic characteristics, functional impairment and physical limitations, concurrent medical conditions, Charlson Comorbidity Index (CCI) scores, and concomitant medication use of patients with OAB; these characteristics are also described by sex and age group (65-74 vs. ≥75 years). We also compare the characteristics of OAB with non-OAB patients. We identified 415 elderly patients with OAB (average age 79 years; 71% female) and 6868 without OAB (average age 77 years; 62% female). Patients with OAB reported high levels of functional impairment as measured by the Activities of Daily Living (44% vs. 33% for non-OAB patients), Instrumental ADL (53% vs. 40% for non-OAB patients), and physical functioning limitation (90% vs. 81% for non-OAB patients) scales. Elderly patients with OAB also experienced high levels of comorbidity burden, as measured by the number of medical conditions (18 vs. 11 for non-OAB patients), CCI (2.1 vs. 1.4 for non-OAB patients), and number of non-OAB-related concomitant medications used (11 vs. 8 for non-OAB patients). Elderly patients with OAB in the general population have high levels of functional impairment and physical limitations, comorbidity, and concomitant medication use. These characteristics should be taken into consideration when managing OAB symptoms and designing future clinical studies. These results, which are representative of elderly patients with OAB in the general US population, should, however, be interpreted in light of the key limitations of the data we used: patients may have been misclassified and medical conditions overestimated due to artifacts of diagnosis coding and our results can only be generalized to patients who were enrolled in Medicare Parts A, B, and D for at least 12 continuous months.


CUL3BPM E3 ubiquitin ligases regulate MYC2, MYC3, and MYC4 stability and JA responses.
The jasmonate (JA)-pathway regulators MYC2, MYC3, and MYC4 are central nodes in plant signaling networks integrating environmental and developmental signals to fine-tune JA defenses and plant growth. Continuous activation of MYC activity is potentially lethal. Hence, MYCs need to be tightly regulated in order to optimize plant fitness. Among the increasing number of mechanisms regulating MYC activity, protein stability is arising as a major player. However, how the levels of MYC proteins are modulated is still poorly understood. Here, we report that MYC2, MYC3, and MYC4 are targets of BPM (BTB/POZ-MATH) proteins, which act as substrate adaptors of CUL3-based E3 ubiquitin ligases. Reduction of function of CUL3BPM in amiR-bpm lines, bpm235 triple mutants, and cul3ab double mutants enhances MYC2 and MYC3 stability and accumulation and potentiates plant responses to JA such as root-growth inhibition and MYC-regulated gene expression. Moreover, MYC3 polyubiquitination levels are reduced in amiR-bpm lines. BPM3 protein is stabilized by JA, suggesting a negative feedback regulatory mechanism to control MYC activity, avoiding harmful runaway responses. Our results uncover a layer for JA-pathway regulation by CUL3BPM-mediated degradation of MYC transcription factors.


An exo-beta-1,3-galactanase having a novel beta-1,3-galactan-binding module from Phanerochaete chrysosporium.
An exo-beta-1,3-galactanase gene from Phanerochaete chrysosporium has been cloned, sequenced, and expressed in Pichia pastoris. The complete amino acid sequence of the exo-beta-1,3-galactanase indicated that the enzyme consists of an N-terminal catalytic module with similarity to glycoside hydrolase family 43 and an additional unknown functional domain similar to carbohydrate-binding module family 6 (CBM6) in the C-terminal region. The molecular mass of the recombinant enzyme was estimated as 55 kDa based on SDS-PAGE. The enzyme showed reactivity only toward beta-1,3-linked galactosyl oligosaccharides and polysaccharide as substrates but did not hydrolyze beta-1,4-linked galacto-oligosaccharides, beta-1,6-linked galacto-oligosaccharides, pectic galactan, larch arabinogalactan, arabinan, gum arabic, debranched arabinan, laminarin, soluble birchwood xylan, or soluble oat spelled xylan. The enzyme also did not hydrolyze beta-1,3-galactosyl galactosaminide, beta-1,3-galactosyl glucosaminide, or beta-1,3-galactosyl arabinofuranoside, suggesting that it specifically cleaves the internal beta-1,3-linkage of two galactosyl residues. High performance liquid chromatographic analysis of the hydrolysis products showed that the enzyme produced galactose from beta-1,3-galactan in an exo-acting manner. However, no activity toward p-nitrophenyl beta-galactopyranoside was detected. When incubated with arabinogalactan proteins, the enzyme produced oligosaccharides together with galactose, suggesting that it is able to bypass beta-1,6-linked galactosyl side chains. The C-terminal CBM6 did not show any affinity for known substrates of CBM6 such as xylan, cellulose, and beta-1,3-glucan, although it bound beta-1,3-galactan when analyzed by affinity electrophoresis. Frontal affinity chromatography for the CBM6 moiety using several kinds of terminal galactose-containing oligosaccharides as the analytes clearly indicated that the CBM6 specifically interacted with oligosaccharides containing a beta-1,3-galactobiose moiety. When the degree of polymerization of galactose oligomers was increased, the binding affinity of the CBM6 showed no marked change.


Long-Chain Fatty Acids Degradation by Desulfomonile Species and Proposal of "Candidatus Desulfomonile Palmitatoxidans".
Microbial communities with the ability to convert long-chain fatty acids (LCFA) coupled to sulfate reduction can be important in the removal of these compounds from wastewater. In this work, an enrichment culture, able to oxidize the long-chain fatty acid palmitate (C16 : 0) coupled to sulfate reduction, was obtained from anaerobic granular sludge. Microscopic analysis of this culture, designated HP culture, revealed that it was mainly composed of one morphotype with a typical collar-like cell wall invagination, a distinct morphological feature of the Desulfomonile genus. 16S rRNA gene amplicon and metagenome-assembled genome (MAG) indeed confirmed that the abundant phylotype in HP culture belong to Desulfomonile genus [ca. 92% 16S rRNA gene sequences closely related to Desulfomonile spp.; and ca. 82% whole genome shotgun (WGS)]. Based on similar cell morphology and average nucleotide identity (ANI) (77%) between the Desulfomonile sp. in HP culture and the type strain Desulfomonile tiedjei strain DCB-1T, we propose a novel species designated as "Candidatus Desulfomonile palmitatoxidans." This bacterium shares 94.3 and 93.6% 16S rRNA gene identity with Desulfomonile limimaris strain DCB-MT and D. tiedjei strain DCB-1T, respectively. Based on sequence abundance of Desulfomonile-morphotype in HP culture, its predominance in the microscopic observations, and presence of several genes coding for enzymes involved in LCFA degradation, the proposed species "Ca. Desulfomonile palmitatoxidans" most probably plays an important role in palmitate degradation in HP culture. Analysis of the growth of HP culture and D. tiedjei strain DCB-1T with short- (butyrate), medium- (caprylate) and long-chain fatty acids (palmitate, stearate, and oleate) showed that both cultures degraded all fatty acids coupled to sulfate reduction, except oleate that was only utilized by HP culture. In the absence of sulfate, neither HP culture, nor D. tiedjei strain DCB-1T degraded palmitate when incubated with Methanobacterium formicicum as a possible methanogenic syntrophic partner. Unlike D. tiedjei strain DCB-1T, "Ca. Desulfomonile palmitatoxidans" lacks reductive dehalogenase genes in its genome, and HP culture was not able to grow by organohalide respiration. An emended description of the genus Desulfomonile is proposed. Our study reveals an unrecognized LCFA degradation feature of the Desulfomonile genus.


Multi-Task Learning for Compositional Data via Sparse Network Lasso.
Multi-task learning is a statistical methodology that aims to improve the generalization performances of estimation and prediction tasks by sharing common information among multiple tasks. On the other hand, compositional data consist of proportions as components summing to one. Because components of compositional data depend on each other, existing methods for multi-task learning cannot be directly applied to them. In the framework of multi-task learning, a network lasso regularization enables us to consider each sample as a single task and construct different models for each one. In this paper, we propose a multi-task learning method for compositional data using a sparse network lasso. We focus on a symmetric form of the log-contrast model, which is a regression model with compositional covariates. Our proposed method enables us to extract latent clusters and relevant variables for compositional data by considering relationships among samples. The effectiveness of the proposed method is evaluated through simulation studies and application to gut microbiome data. Both results show that the prediction accuracy of our proposed method is better than existing methods when information about relationships among samples is appropriately obtained.


Genetic variation within a neutralizing domain on the haemagglutinin-neuraminidase glycoprotein of Newcastle disease virus.
Previously, a panel of monoclonal antibodies recognizing epitopes in four antigenic sites on the haemagglutinin-neuraminidase (HN) glycoprotein of the Australia-Victoria strain of Newcastle disease virus were used in strain comparisons. Epitopes in three sites were found to be conserved while the epitope recognized by the single antibody to site 3 was not. A new panel of antibodies is described, two of which bind to epitopes in site 3 and six of which bind to a site (site 1,4) that overlaps with sites 1 and 4 as determined by analyses of variants, temperature-sensitive mutants, and strains by assays of neutralization of infectivity and binding in a radioimmunoassay. Neutralization of heterologous strains with the panel of antibodies revealed that both new site 3 epitopes are also highly divergent, while three additional epitopes outside site 3 (those in site 1,4) are highly conserved. The new site 3 antibodies can bind to virions of several heterologous strains without neutralizing infectivity. Thus, of the 10 epitopes we have now examined, all of three in site 3 are specific with respect to neutralization of infectivity for the homologous strain, while all of seven in other sites are conserved in heterologous strains. This suggests that the strain specificity originally described for a single site 3 epitope is, instead, a property of a much more extensive, poorly conserved domain on the HN molecule.


Cerebellar guidance of premotor network development and sensorimotor learning.
Single unit and imaging studies have shown that the cerebellum is especially active during the acquisition phase of certain motor and cognitive tasks. These data are consistent with the hypothesis that particular sensorimotor procedures are acquired and stored in the cerebellar cortex and that this knowledge can then be exported to the cerebral cortex and premotor networks for more efficient execution. In this article we present a model to illustrate how the cerebellar cortex might guide the development of cortical-cerebellar network connections and how a similar mechanism operating in the adult could mediate the exportation of sensorimotor knowledge from the cerebellum to the motor cortex. The model consists of a three-layered recurrent network representing the cerebello-thalamocortical-ponto-cerebellar limb premotor network. The cerebellar cortex is not explicitly modeled. Our simulations show that Hebbian learning combined with weight normalization allows the emergence of reciprocal and modular structure in the limb premotor network. Reciprocal connections allow activity to reverberate around specific loops. Modularity organizes the connections into specific channels. Furthermore, we show that cerebellar learning can be exported to motor cortex through these modular and reciprocal premotor circuits. In particular, we simulate developmental alignment of visuomotor relations and their realignment as a consequence of prism exposure. The exportation of sensorimotor knowledge from the cerebellum to the motor cortex may allow faster and more efficient execution of learned motor responses.


Characterization of human transcription factor function and patterns of gene regulation in HepG2 cells.
Transcription factors (TFs) are trans-acting proteins that bind cis-regulatory elements (CREs) in DNA to control gene expression. Here, we analyzed the genomic localization profiles of 529 sequence-specific TFs and 151 cofactors and chromatin regulators in the human cancer cell line HepG2, for a total of 680 broadly termed DNA-associated proteins (DAPs). We used this deep collection to model each TF's impact on gene expression, and identified a cohort of 26 candidate transcriptional repressors. We examine high occupancy target (HOT) sites in the context of three-dimensional genome organization and show biased motif placement in distal-promoter connections involving HOT sites. We also found a substantial number of closed chromatin regions with multiple DAPs bound, and explored their properties, finding that a MAFF/MAFK TF pair correlates with transcriptional repression. Altogether, these analyses provide novel insights into the regulatory logic of the human cell line HepG2 genome and show the usefulness of large genomic analyses for elucidation of individual TF functions.


First record of entomopathogenic nematodes (Steinernematidae and Heterorhabditidae) in Costa Rica.
A survey of entomopathogenic nematodes was conducted in the north Pacific (Guanacaste Conservation Area) and southeast Caribbean (Gandoca-Manzanillo Natural Refuge) regions of Costa Rica. Out of a total of 41 soil samples, 5 were positive for entomopathogenic nematodes (20.5%), with 3 (12.3%) containing Steinernema and 2 (8.2%) Heterorhabditis isolates. Morphological and molecular studies were undertaken to characterize these isolates. The Heterorhabditis isolates were identified as Heterorhabditis indica and the three Steinernema isolates were identified as two new undescribed species. H. indica was recovered from a coastal dry forest. Steinernema n. sp. 1 was isolated from a rainforest valley, between volcanoes. Steinernema sp. n. 2 was isolated from sand dunes in the Caribbean Coast (Punta Uva) near the rainforest strip along the coast. Although limited to two geographic regions, this study suggests entomopathogenic nematodes may be diverse and perhaps widely distributed in Costa Rica. A more intensive survey, covering all geographic regions is currently undergoing.


MAPT (Tau) expression is a biomarker for an increased rate of survival for low‑grade glioma.
The association of MAPT (Tau) with various tauopathies and other neurological disorders has long been established. However, the role of microtubule‑associated protein Tau (MAPT) expression in brain cancer is largely unknown. To determine whether MAPT expression is related to low‑grade glioma (LGG) survival rates, RNASeq data representing samples from the Cancer Genome Atlas (TCGA) were assessed. Results revealed that high expression of the MAPT gene is very strongly associated with increased overall and disease‑free survival in LGG but not in breast cancer or melanoma. No such association was apparent for either amyloid precursor protein or α‑synuclein gene expression. The expression levels of particular apoptosis‑ and pro‑proliferative‑effector genes were consistent with the Tau‑associated increased survival rates. It has been well established that the Tau protein plays a neurodegenerative role, and in this study we identified, for the first time, a potential cell apoptosis function that Tau may play in cancers of the central nervous system.


A white blood cell index as the main prognostic factor in t(8;21) acute myeloid leukemia (AML): a survey of 161 cases from the French AML Intergroup.
While the t(8;21) translocation is one of the most recurrent chromosomal abnormalities in acute myeloid leukemia, prognostic studies have been hampered by the relatively few number of patients reported. We thus performed a large retrospective study in 161 adults and children with t(8;21) acute myeloid leukemia, all prospectively enrolled in 6 different trials conducted in France between 1987 and 1998 (median follow-up 4.9 years). Prognostic studies were performed in the 154 patients who achieved a complete remission. Individual data were registered, including sex, age, blood and marrow counts, extramedullary disease, and cytogenetics. The value of allogeneic stem cell transplantation versus chemotherapy as postremission therapy was evaluated according to the intent-to-treat principle. Estimated 5-year disease-free survival (DFS) and overall survival were 52% and 59%, respectively. Outcome was not significantly better in patients from the stem cell transplantation group (estimated 5-year DFS and survival, 56% vs 52% and 67% vs 57%; P =.55 and.64, respectively). White blood cell count (WBC) was the only identified prognostic factor. To further take into account the spontaneous differentiation potential of the leukemic clone, a WBC index was derived as the product of WBC by the ratio of marrow blast. This WBC index was a more powerful factor than the original WBC, allowing us to distinguish 3 subgroups of patients with different outcomes (low index, < 2.5; intermediate index, 2.5-20; high index, 20 or more). In multivariate analysis, the WBC index was the only prognostic factor for DFS (P =.003), complete remission duration (P =.002), and overall survival (P =.04).


Vitamin D Receptor: A Novel Therapeutic Target for Kidney Diseases.
Kidney disease is a serious problem that adversely affects human health, but critical knowledge is lacking on how to effectively treat established chronic kidney disease. Mounting evidence from animal and clinical studies has suggested that Vitamin D Receptor (VDR) activation has beneficial effects on various renal diseases. A structured search of published research literature regarding VDR structure and function, VDR in various renal diseases (e.g., IgA nephropathy, idiopathic nephrotic syndrome, renal cell carcinoma, diabetic nephropathy, lupus nephritis) and therapies targeting VDR was performed for several databases. Included in this study are the results from 177 published research articles. Evidence from these papers indicates that VDR activation is involved in the protection against renal injury in kidney diseases by a variety of mechanisms, including suppression of RAS activation, anti-inflammation, inhibiting renal fibrogenesis, restoring mitochondrial function, suppression of autoimmunity and renal cell apoptosis. VDR offers an attractive druggable target for renal diseases. Increasing our understanding of VDR in the kidney is a fertile area of research and may provide effective weapons in the fight against kidney diseases.


[Isolation of new alleles of the swine TLR4 gene and analysis of its genetic variation].
Using the PCR-SSCP method, the genetic variation in exon 1 of the TLR4 gene was detected among 893 animals, including Asian wild boars, 3 imported commercial and 10 Chinese indigenous swine breeds. This was conducted to analyze the polymorphisms of exon 1 of TLR4 gene in native and foreign pig breeds and aimed at providing a theoretical foundation for further research on the role that TLR4 gene played in immune and defense system. New alleles were isolated for exon 1 of the swine TLR4 gene for the first time, There were 6 genotypes and 3 alleles, in which the Duroc appeared AA, BB, CC, AB, AC and BC genotypes; Sutai pig, which has Duroc pig origin, were detected to be BB, CC, and BC genotypes; Yorkshire and Landrace were detected to be CC and BC genotypes. Wild boar and all 10 Chinese native pig breeds appeared highly conserved in exon 1 of TLR4 gene, with only CC genotype. Among the 3 homozygous genotypes, the CC genotype matches the sequence in GenBank, while a G93C synonymous mutation and a G194A nonsense mutation were found in the BB and AA genotypes, respectively. The correlation between these two mutation points of TLR4 gene with resistance to stress and disease is worthy of further study.


APOE ε4/ε4 diminishes neurotrophic function of human iPSC-derived astrocytes.
The ε4 allele of the APOE gene encoding apolipoprotein E (apoE) is a strong genetic risk factor for aging-related cognitive decline as well as late-onset Alzheimer's disease (AD) compared to the common ε3 allele. In the central nervous system, apoE is produced primarily by astrocytes and functions in transporting lipids including cholesterol to support neuronal homeostasis and synaptic integrity. Although mouse models and corresponding primary cells have provided valuable tools for studying apoE isoform-dependent functions, recent studies have shown that human astrocytes have a distinct gene expression profile compare with rodent astrocytes. Human induced pluripotent stem cells (iPSCs) derived from individuals carrying specific gene variants or mutations provide an alternative cellular model more relevant to humans upon differentiation into specific cell types. Thus, we reprogramed human skin fibroblasts from cognitively normal individuals carrying APOE ε3/ε3 or ε4/ε4 genotype to iPSC clones and further differentiated them into neural progenitor cells and then astrocytes. We found that human iPSC-derived astrocytes secreted abundant apoE with apoE4 lipoprotein particles less lipidated compared to apoE3 particles. More importantly, human iPSC-derived astrocytes were capable of promoting neuronal survival and synaptogenesis when co-cultured with iPSC-derived neurons with APOE ε4/ε4 astrocytes less effective in supporting these neurotrophic functions than those with APOE ε3/ε3 genotype. Taken together, our findings demonstrate APOE genotype-dependent effects using human iPSC-derived astrocytes and provide novel evidence that the human iPSC-based model system is a strong tool to explore how apoE isoforms contribute to neurodegenerative diseases.


Tension Strain-Softening and Compression Strain-Stiffening Behavior of Brain White Matter.
Brain, the most important component of the central nervous system (CNS), is a soft tissue with a complex structure. Understanding the role of brain tissue microstructure in mechanical properties is essential to have a more profound knowledge of how brain development, disease, and injury occur. While many studies have investigated the mechanical behavior of brain tissue under various loading conditions, there has not been a clear explanation for variation reported for material properties of brain tissue. The current study compares the ex-vivo mechanical properties of brain tissue under two loading modes, namely compression and tension, and aims to explain the differences observed by closely examining the microstructure under loading. We tested bovine brain samples under uniaxial tension and compression loading conditions, and fitted hyperelastic material parameters. At 20% strain, we observed that the shear modulus of brain tissue in compression is about 6 times higher than in tension. In addition, we observed that brain tissue exhibited strain-stiffening in compression and strain-softening in tension. In order to investigate the effect of loading modes on the tissue microstructure, we fixed the samples using a novel method that enabled keeping the samples at the loaded stage during the fixation process. Based on the results of histology, we hypothesize that during compressive loading, the strain-stiffening behavior of the tissue could be attributed to glial cell bodies being pushed against surroundings, contacting each other and resisting compression, while during tension, cell connections are detached and the tissue displays softening behavior.


Analysis of the antitumoral mechanisms of lipopolysaccharide against glioblastoma multiforme.
Our objective was to analyze the lipopolysaccharide (LPS) antitumoral effect upon glioblastoma, including whether the lipid A subunit alone can elicit glioblastoma regression, whether dexamethasone suppresses this response to LPS, whether B and T lymphocytes factor in this response, and whether this antitumoral effect of LPS provides resistance against subsequent challenge with glioblastoma. Mice (BALB/c, nude or SCID) implanted with s.c. DBT glioblastomas were treated with LPS (with or without dexamethasone) or with lipid A. A subset of BALB/c mice in which s.c. DBT glioblastomas had previously been eradicated using LPS were re-implanted with s.c. or intracranial (i.c.) DBT cells. For mice with s.c. tumors, mean tumor masses (MTM) were compared between groups. Survival was compared for mice with i.c. tumors. Lipid A caused near complete tumor regression of DBT glioblastomas in BALB/c mice (p<0.0001). Dexamethasone did not alter the antitumoral effect of LPS (p=0.48). LPS reduced the MTM of s.c. glioblastomas in T lymphocyte-deficient nude mice, but not as effectively as in immunocompetent mice. The antitumoral response to LPS for T and B lymphocyte-deficient SCID mice bearing DBT glioblastomas was similar to that for nude mice. Eradication of s.c. DBT glioblastoma in BALB/c provided partial resistance to subsequent challenge with s.c. or i.c. glioblastoma. We conclude that the LPS-mediated antitumoral response against glioblastoma is dependent upon the lipid A subunit of LPS, partially dependent upon T lymphocytes, independent of B lymphocytes, unaffected by dexamethasone and provides partial protection against subsequent challenges with glioblastoma.


Comparison of pressure distribution under a conventional saddle and a treeless saddle at sitting trot.
It can be a challenge to find a conventional saddle that is a good fit for both horse and rider. An increasing number of riders are purchasing treeless saddles because they are thought to fit a wider range of equine back shapes, but there is only limited research to support this theory. The objective of this study was to compare the total force and pressure distribution patterns on the horse's back with conventional and treeless saddles. The experimental hypotheses were that the conventional saddle would distribute the force over a larger area with lower mean and maximal pressures than the treeless saddle. Eight horses were ridden by a single rider at sitting trot with conventional and treeless saddles. An electronic pressure mat measured total force, area of saddle contact, maximal pressure and area with mean pressure >11 kPa for 10 strides with each saddle. Univariate ANOVA (P<0.05) was used to detect differences between saddles. Compared with the treeless saddle, the conventional saddle distributed the rider's bodyweight over a larger area, had lower mean and maximal pressures and fewer sensors recording mean pressure >11 kPa. These findings suggested that the saddle tree was effective in distributing the weight of the saddle and rider over a larger area and in avoiding localized areas of force concentration.


Self-reported Symptoms of Depression Among Chinese Rural-to-Urban Migrants and Left-Behind Family Members.
There were an estimated 247 million rural-to-urban migrant workers in China in 2016, yet at a national level, there is scant evidence on the association of migration with mental health among migrants and their left-behind family members. To examine the association of rural-to-urban migration with symptoms of depression among migrants and left-behind family members aged 45 years and older. Using representative cross-sectional data of 14 332 middle-aged and older adults from the 2015 China Health and Retirement Longitudinal Survey, regression analyses were conducted to examine the association of depressive symptoms with rural-to-urban migration status in urban areas and the association of depressive symptoms with left-behind status in rural areas. The statistical analysis was performed from January to August 2018. Migration status (defined as having a rural hukou [household registration record]) in urban areas and left-behind status (defined as having a spouse or child living in another area) in rural areas. Depressive symptoms measured on the 10-item Center for Epidemiological Studies Depression (CES-D-10) scale. A total of 14 332 middle-aged and elderly participants (mean [SD] age, 59.84 [9.51] years; 7394 [51.6%] women) were included, of whom 4404 (30.7%) lived in urban areas and 9928 (69.3%) lived in rural areas. In urban areas, 1607 participants (36.2%) were rural-to-urban migrants, and the remaining 2797 participants (72.8%) were local residents. In rural areas, 3405 participants (34.3%) were left-behind family members, and the remaining 6523 participants (65.7%) were not. Compared with urban residents, rural-to-urban migrants had higher CES-D-10 scores after adjustment for covariates (β = 0.74; 95% CI, 0.08-1.40; P = .03; standard errors clustered at the household level henceforth). Compared with intact-family rural residents, left-behind spouses had higher CES-D-10 scores after adjustment for covariates (β = 0.54; 95% CI, 0.05-1.03; P = .03). Rural-to-urban migration in China was associated with poor mental health for migrants and their left-behind spouses. Short-term policies, such as building community social facilities, may prove effective, but long-term solutions should address issues related to economic and social exclusions and the lack of a social security system in rural China.


Unique small molecule entry inhibitors of hemorrhagic fever arenaviruses.
Viral hemorrhagic fevers caused by the arenaviruses Lassa virus in Africa and Machupo, Guanarito, Junin, and Sabia virus in South America are among the most devastating emerging human diseases with fatality rates of 15-35% and a limited antiviral therapeutic repertoire available. Here we used high throughput screening of synthetic combinatorial small molecule libraries to identify inhibitors of arenavirus infection using pseudotyped virion particles bearing the glycoproteins (GPs) of highly pathogenic arenaviruses. Our screening efforts resulted in the discovery of a series of novel small molecule inhibitors of viral entry that are highly active against both Old World and New World hemorrhagic arenaviruses. We observed potent inhibition of infection of human and primate cells with live hemorrhagic arenaviruses (IC(50)=500-800 nm). Investigations of the mechanism of action revealed that the candidate compounds efficiently block pH-dependent fusion by the arenavirus GPs (IC(50) of 200-350 nm). Although our lead compounds were potent against phylogenetically distant arenaviruses, they did not show activity against other enveloped viruses with class I viral fusion proteins, indicating specificity for arenavirus GP-mediated membrane fusion.


Diagnosis and management of microscopic colitis.
To describe microscopic colitis (MC) and a stepwise approach to its diagnosis and management. MEDLINE was searched from January 1996 to August 2002. Controlled trials were sought, but due to their relative rarity, most articles cited are well designed retrospective studies and reviews. Microscopic colitis in both its collagenous and lymphocytic forms is a relatively common and important cause of diarrhea in middle-aged and elderly patients. It usually presents as chronic watery diarrhea. Results of endoscopic and radiologic examinations are normal; specific histologic findings are seen on colonic biopsy. Once the diagnosis is confirmed, we suggest a stepwise approach to therapy. First-line therapies include antidiarrheal agents; second-line include bismuth subsalicylates and budesonide; third-line include cholestyramine and 5-aminosalicylic acid agents. When patients fail to respond to these therapies, oral corticosteroids or immune modulatory therapy should be considered. For severe unresponsive cases, colectomy is the last resort. Microscopic colitis is an important cause of chronic diarrhea. Once the diagnosis is confirmed, a stepwise approach to treatment is suggested.


Automated sample preparation in a microfluidic culture device for cellular metabolomics.
Sample pretreatment in conventional cellular metabolomics entails rigorous lysis and extraction steps which increase the duration as well as limit the consistency of these experiments. We report a biomimetic cell culture microfluidic device (MFD) which is coupled with an automated system for rapid, reproducible cell lysis using a combination of electrical and chemical mechanisms. In-channel microelectrodes were created using facile fabrication methods, enabling the application of electric fields up to 1000 V cm(-1). Using this platform, average lysing times were 7.12 s and 3.03 s for chips with no electric fields and electric fields above 200 V cm(-1), respectively. Overall, the electroporation MFDs yielded a ∼10-fold improvement in lysing time over standard chemical approaches. Detection of multiple intracellular nucleotides and energy metabolites in MFD lysates was demonstrated using two different MS platforms. This work will allow for the integrated culture, automated lysis, and metabolic analysis of cells in an MFD which doubles as a biomimetic model of the vasculature.


Associations of Cardiac, Kidney, and Diabetes Biomarkers With Peripheral Neuropathy among Older Adults in the Atherosclerosis Risk in Communities (ARIC) Study.
The aim of this study was to assess the association of high-sensitivity cardiac troponin (hs-cTnT) and other cardiac, kidney, hyperglycemia, and inflammatory biomarkers with peripheral neuropathy (PN) in a community-based population. We conducted a cross-sectional analysis of 3056 black and white participants in the Atherosclerosis Risk in Communities (ARIC) study who underwent standardized monofilament PN testing and had measures of cardiac function (hs-cTnT, N-terminal pro-B-type natriuretic peptide [NT-proBNP], and growth differentiation factor 15 [GDF15]), kidney function (serum creatinine, cystatin C, β-2 microglobulin, urine albumin-to-creatinine ratio), hyperglycemia (fasting glucose, hemoglobin A1c [Hb A1c], fructosamine, glycated albumin, 1,5-anhydroglucitol), and inflammation (C-reactive protein) assessed at visit 6 (2016-2017; age 71-94 years). We used logistic regression to assess the associations of these biomarkers (modeled in diabetes-specific tertiles) with PN in older adults with and without diabetes after adjusting for traditional risk factors. In total, 33.5% of participants had PN (37.3% with diabetes and 31.9% without diabetes). There was an independent association of hs-cTnT with PN regardless of diabetes status (diabetes T3 vs. T1: odds ratio [OR], 2.15 [95% CI, 1.44-3.22]; no diabetes: OR, 2.31 [95%CI, 1.76-3.03]; P = 0.72 for interaction). Among participants without diabetes, there were also significant associations of NT-proBNP (OR, 1.40 [95% CI, 1.08-1.81]) and urine albumin-to-creatinine ratio (OR, 1.55 [95% CI, 1.22-1.97]) with PN. Associations of hyperglycemia biomarkers including Hb A1c (OR, 1.76 [95% CI, 1.22-2.54]), fructosamine (OR, 1.71 [95% CI, 1.19-2.46]), and glycated albumin (OR, 1.45 [95% CI, 1.03-2.03]) with PN were significant only among participants with diabetes. Overall, hs-cTnT appears to be a global marker of end organ damage, including PN. Laboratory biomarkers may be able to help us identify those individuals with PN.


Selective review of cognitive aging.
Research concerned with relations between adult age and cognitive functioning is briefly reviewed. The coverage is necessarily selective, and is organized in terms of five major questions. These are what abilities are related to age, how many distinct influences are contributing to the relations between age and cognitive functioning, do the differences between people increase with advancing age, what is responsible for the discrepancies between cross-sectional and longitudinal age comparisons of cognitive functioning, and what methods can be used to identify causes of age-related influences on cognition. Although definitive answers are not yet possible, quite a bit of information relevant to the questions is now available. Moreover, the existing information has implications for the design, analysis, and interpretation of cognitive and neuropsychological research concerned with aging.


Chitin nanofibers prepared by enzymatic hydrolysis: Characterization and application for Pickering emulsions.
Chitin nanofibers (ChNFs) have a wide range of applications in numerous fields owing to their exceptional material properties and biological functionality. This research focused on producing ChNFs with diameters of 20-70 nm using chitinase and ultrasound from crayfish shells. The impact of enzymatic duration on ChNF yield and performance was investigated. Results revealed ChNFs forming a high aspect ratio network structure. Chitinase hydrolysis enhanced ChNF dispersion and yield while improving crystallinity and thermal stability without significantly altering their chemical structure. Enzymatically modified ChNF suspensions also exhibited stable rheological properties. Moreover, ChNFs showed good emulsification and emulsion stability in Pickering emulsion. The mechanism may be the effective adsorption of ChNFs at the oil-water interface, and the formation of a ChNF network in the continuous phase that prevents droplet coalescence. This study highlights that the potential of chitinase and ultrasound for the production of ChNFs and the utilization of crayfish shell waste.


Culture and characterization of various porcine integumentary-connective tissue-derived mesenchymal stromal cells to facilitate tissue adhesion to percutaneous metal implants.
Transdermal osseointegrated prosthesis have relatively high infection rates leading to implant revision or failure. A principle cause for this complication is the absence of a durable impervious biomechanical seal at the interface of the hard structure (implant) and adjacent soft tissues. This study explores the possibility of recapitulating an analogous cellular musculoskeletal-connective tissue interface, which is present at naturally occurring integumentary tissues where a hard structure exits the skin, such as the nail bed, hoof, and tooth. Porcine mesenchymal stromal cells (pMSCs) were derived from nine different porcine integumentary and connective tissues: hoof-associated superficial flexor tendon, molar-associated periodontal ligament, Achilles tendon, adipose tissue and skin dermis from the hind limb and abdominal regions, bone marrow and muscle. For all nine pMSCs, the phenotype, multi-lineage differentiation potential and their adhesiveness to clinical grade titanium was characterized. Transcriptomic analysis of 11 common genes encoding cytoskeletal proteins VIM (Vimentin), cell-cell and cell-matrix adhesion genes (Vinculin, Integrin β1, Integrin β2, CD9, CD151), and for ECM genes (Collagen-1a1, Collagen-4a1, Fibronectin, Laminin-α5, Contactin-3) in early passaged cells was performed using qRT-PCR. All tissue-derived pMSCs were characterized as mesenchymal origin by adherence to plastic, expression of cell surface markers including CD29, CD44, CD90, and CD105, and lack of hematopoietic (CD11b) and endothelial (CD31) markers. All pMSCs differentiated into osteoblasts, adipocytes and chondrocytes, albeit at varying degrees, under specific culture conditions. Among the eleven adhesion genes evaluated, the cytoskeletal intermediate filament vimentin was found highly expressed in pMSC isolated from all tissues, followed by genes for the extracellular matrix proteins Fibronectin and Collagen-1a1. Expression of Vimentin was the highest in Achilles tendon, while Fibronectin and Col1agen-1a1 were highest in molar and hoof-associated superficial flexor tendon bone marrow, respectively. Achilles tendon ranked the highest in both multilineage differentiation and adhesion assessments to titanium metal. These findings support further preclinical research of these tissue specific-derived MSCs in vivo in a transdermal osseointegration implant model.


Cost of conducting Measles-Rubella vaccination campaign in India.
A Measles-Rubella (MR) vaccination campaign was launched in India in a phased manner in February 2017 to cover children aged 9 months to 15 years. As evidence on campaign vaccine delivery costs is limited, the delivery cost for MR campaign from a government provider perspective was estimated in four Indian states, namely, Assam, Gujarat, Himachal Pradesh, and Uttar Pradesh. Costs were calculated in top-down and bottom-up approaches using data collected from 84 sites at different administrative levels and immunization partners in the study states from August 2019 to March 2020. All costs were presented in 2019 US$ and Indian Rupee (INR). The financial cost per dose of the MR campaign including all partner support ranged from US$0.16 (INR 10.95) in Uttar Pradesh to US$0.34 (INR 24.13) in Gujarat. In Uttar Pradesh, the full economic cost per dose was US$0.87 (INR 61.39). The key financial cost drivers were incentives related to service delivery and supervision, the printing of reporting formats for record-keeping, social mobilization, and advocacy. The financial delivery cost per dose estimated was higher than the government pre-fixed budget per child for the MR campaign, probably indicating an insufficient budget. However, the study found underutilization of MR budget in two states and use of other sources of funding for the campaign indicating the need for proper utilization of the campaign budgets by the states. Unit cost information generated from this study will be useful for planning, cost projections, and economic analysis of future vaccination campaigns in India.


Effect of Microbiota in the Development of Breast Cancer.
Breast cancer is the most frequent cancer among women and causes the greatest number of cancer-related death among women all over the world. It approximately accounts for 15% of all cancer death. The human microbiota is the term applied to the aggregate of microbes that live in different habitats of living organisms 'bodies, including the gut, skin, vagina, and mouth, as well as nose, conjunctiva, pharynx, and urethra, among others. Increasing evidence is pointing to the role of the microbiome in the occurrence and development of a variety of cancers. Intestinal microbiome imbalance is related to the occurrence of gastrointestinal tumors, such as esophageal, gastric, colorectal, and gallbladder cancer. The present study aimed to identify the role of microbiota in the development of breast cancer. The women with breast cancer (n=130) in this study were in the age range of 25-75 years. The study was conducted in Kirkuk city of Iraq from September 10, 2019, to March 15, 2020. The control group included 20 women diagnosed with benign breast lesions in the age range 25-75 years, who matched the women in the patient group. Blood samples and breast tissue samples were taken from patients with breast cancer and benign breast lesions. Blood samples were examined through immunological methods, enzyme-linked immunosorbent assay (ELISA) was adopted for the detection of interleukin-19 (IL-19). Breast tissue samples were taken from breast cancer and benign breast lesions patients to isolate and identify bacteria. Based on the obtained results, only 6 out of 30 (20%) cultured breast tissue samples from women with breast cancer showed bacterial growth. In total, 4 (67%) and 2(33%) of these 6 positive cultures were Escherichia coli was and Staphylococcus aureus, respectively, and this relation was statistically significant. However, no bacterial growth was observed on the cultured breast tissue samples taken from women with benign breast lesions. Moreover, the difference between women with a positive and negative result of bacterial culture and stages of breast cancer was statistically non-significant. It is worth mentioning that 50 % of women with breast cancer and bacterial growth were within the age range of 40-49 year. The present study revealed that the difference between women with breast cancer and those with benign breast lesions was statistically highly significant according to the place of residence. In addition, the mean level of IL-19 among women with breast cancer was lower than that in women with benign breast lesions, and this relation was statistically highly significant.


Protein corona-resistant SERS tags for live cell detection of integrin receptors.
Chemical signals are conveyed to cells through ligand-receptor binding, triggering cascades of biochemical reactions and resulting in pivotal cellular functions. These binding events are important in understanding membrane signaling and drug interactions. To probe ligand-receptor binding, surface enhanced Raman scattering (SERS) tags are a promising tool. SERS tags are plasmonic nanostructures functionalized with a protective coating, a Raman reporter molecule, and a biorecognition element. In biological fluids, native proteins have affinity for bare nanoparticles and form a protein corona. SERS tags have a protective shell which eliminates this complication. It is important to analyze ligand-receptor binding with SERS tags in live cells since cell fixatives alter protein structure, leading to spectral changes and data misinterpretation. In this study, we synthesized a novel SERS tag by creating a mixed monolayer of the small cyclic arginine-glycine-aspartic acid-phenylalanine-cysteine (RGDFC) peptide and 4-mercaptobenzonitrile (MBN) on the surface of spherical gold nanoparticles (Au NP). Au-RGDFC-MBN NP showed resistance to PC formation and were successfully detected in both fixed and living human metastatic colon cancer cells.


Expression levels of enzymes generating NO and CO in islets of murine and human diabetes.
The possible implication of the gasotransmitters NO and CO for the development of diabetes remains unresolved. Our previous investigations in rodents suggested NO being inhibitory, and CO stimulatory, to glucose-stimulated insulin secretion (GSIS). Here we studied the possible role of these gasotransmitters in both murine and human type 2 diabetes (T2D) by mapping the expression pattern of neural nitric oxide synthase (nNOS), inducible NOS (iNOS), constitutive heme oxygenase (HO-2), and inducible HO (HO-1) in isolated pancreatic islets. Two variants of obese murine diabetes with distinct phenotype, the db/db and the ob/ob mouse, were studied at the initiation of the diabetic condition. Plasma glucose and plasma insulin were recorded and β-cell expression levels of the different enzymes were measured with confocal microscopy and fluorescence intensity recordings. In human islets taken from nondiabetic controls (ND) and type 2 diabetes (T2D) the expression of the enzymes was analyzed by RNA-sequencing and qPCR. At the initiation of murine diabetes plasma glucose was slightly increased, whereas plasma insulin was extremely enhanced in both db/db and ob/ob mice. The β-cell expression of nNOS and iNOS was markedly increased over controls in db/db mice, known to develop severe diabetes, while it was very low in ob/ob mice, known to develop mild diabetes. HO-2 expression was unaffected in db/db and modestly decreased in ob/ob mice. HO-1 expression was slightly enhanced in ob/ob, but, in contrast, extremely enhanced in db/db mice, suggesting a counteracting, antidiabetic action by CO. Moreover, the diabetic pattern of highly increased nNOS, iNOS and HO-1 expression seen in db/db mice was also fully recognized in human T2D islets. These results suggest that increased expression of the NOS-enzymes, especially an early upregulation of nNOS, could be involved in the initial development of the severe diabetes of db/db mice as well as in human T2D. Hence, nNOS, iNOS and HO-1 might be regarded as interesting targets to take into consideration in the early treatment of a diabetic condition in different variants of T2D.


Emerging health issues: the widening challenge for population health promotion.
The spectrum of tasks for health promotion has widened since the Ottawa Charter was signed. In 1986, infectious diseases still seemed in retreat, the potential extent of HIV/AIDS was unrecognized, the Green Revolution was at its height and global poverty appeared less intractable. Global climate change had not yet emerged as a major threat to development and health. Most economists forecast continuous improvement, and chronic diseases were broadly anticipated as the next major health issue. Today, although many broadly averaged measures of population health have improved, many of the determinants of global health have faltered. Many infectious diseases have emerged; others have unexpectedly reappeared. Reasons include urban crowding, environmental changes, altered sexual relations, intensified food production and increased mobility and trade. Foremost, however, is the persistence of poverty and the exacerbation of regional and global inequality. Life expectancy has unexpectedly declined in several countries. Rather than being a faint echo from an earlier time of hardship, these declines could signify the future. Relatedly, the demographic and epidemiological transitions have faltered. In some regions, declining fertility has overshot that needed for optimal age structure, whereas elsewhere mortality increases have reduced population growth rates, despite continuing high fertility. Few, if any, Millennium Development Goals (MDG), including those for health and sustainability, seem achievable. Policy-makers generally misunderstand the link between environmental sustainability (MDG #7) and health. Many health workers also fail to realize that social cohesion and sustainability--maintenance of the Earth's ecological and geophysical systems--is a necessary basis for health. In sum, these issues present an enormous challenge to health. Health promotion must address population health influences that transcend national boundaries and generations and engage with the development, human rights and environmental movements. The big task is to promote sustainable environmental and social conditions that bring enduring and equitable health gains.


Increased fibrosis and impaired intratumoral accumulation of macromolecules in a murine model of pancreatic cancer co-administered with FGF-2.
Pancreatic cancer is notorious for its poor prognosis. The histopathologic characteristic of pancreatic ductal adenocarcinoma (PDAC), which is the most common type of pancreatic cancer, is fibrosis within tumor tissue. Although fibrosis within tumor tissue is thought to impede drug therapy by interfering with the intratumoral accumulation of anti-tumor drugs, this hypothesis has yet to be proven directly in preclinical models. Here, we evaluated the effect of enhanced fibrosis on intratumoral accumulation of macromolecular drugs by increasing fibrosis in a murine tumor model of subcutaneously xenografted BxPC-3, a human PDAC cell line. When fibroblast growth factor-2 (FGF-2) was co-administered upon BxPC-3 inoculation, stromal fibrotic area was increased and was characterized by augmented murine collagen accumulation compared to inoculation of BxPC-3 alone, which correlated with increased monocyte/macrophage contents in the tumor tissues. We further discovered that the intratumoral accumulation of intravenously administrated fluorescein isothiocyanate-dextran of 2,000,000Da (2MDa) was significantly reduced in the FGF-2 co-administered tumors despite unaltered hyaluronan accumulation and pericyte coverage of the tumor neovasculature and increased lymphangiogenesis. Finally, we found that FGF-2 co-administered tumors are more refractory to macromolecular drug therapy using nab-paclitaxel (Abraxane). The model established and analyzed in this study, characterized by increased fibrotic component, provides a preclinical animal model suited to predict the intratumoral accumulation of macromolecular drugs and to evaluate the efficacy of drugs targeting the tumor stroma.


Comparison of Nd:YAG and diode laser irradiation during intracoronal bleaching with sodium perborate: color and Raman spectroscopy analysis.
The aim of this study was to evaluate the color and enamel structure changes after intracoronal bleaching with sodium perborate under neodymium:yttrium-aluminum-garnet (Nd:YAG) and diode laser irradiation. Although some studies investigated the efficacy of laser irradiation during intracoronal bleaching, no study has been conducted to investigate the changes in both color and enamel structure during intracoronal bleaching with laser irradiation. Thirty-six extracted mandibular incisors were used. The root canals were prepared and filled with AH Plus and gutta-percha. Baseline color values and Raman spectra were obtained from all samples. The samples were randomly divided into three groups as follows: group 1, intracoronal bleaching with sodium perborate plus NdYAG laser irradiation; group 2, intracoronal bleaching with sodium perborate plus diode laser irradiation; and group 3, intracoronal bleaching with sodium perborate without any laser irradiation. Following the bleaching procedures, the final baseline color values and Raman spectra were obtained. The baseline and final values were statistically analyzed. Statistical analyses were performed with SPSS 18.0 software using Anova and the Kruskal-Wallis test. There was a significant difference between the Nd:YAG laser and control groups (p<0.05). There was no significant difference between the diode laser group and the control group, or the diode laser group and the Nd:YAG laser group. There was no significant difference among all groups in terms of Raman relative intensity (RRI) and fluorescence intensity (FI) percentage values (p>0.05). Laser application, especially Nd:YAG laser irradiation, was able to increase the efficacy of intracoronal bleaching with sodium perborate. Laser irradiation significantly increased the bleaching efficacy without any changes in the enamel surface structure.


Reovirus sigma NS protein localizes to inclusions through an association requiring the mu NS amino terminus.
Cells infected with mammalian reoviruses contain phase-dense inclusions, called viral factories, in which viral replication and assembly are thought to occur. The major reovirus nonstructural protein mu NS forms morphologically similar phase-dense inclusions when expressed in the absence of other viral proteins, suggesting it is a primary determinant of factory formation. In this study we examined the localization of the other major reovirus nonstructural protein, sigma NS. Although sigma NS colocalized with mu NS in viral factories during infection, it was distributed diffusely throughout the cell when expressed in the absence of mu NS. When coexpressed with mu NS, sigma NS was redistributed and colocalized with mu NS inclusions, indicating that the two proteins associate in the absence of other viral proteins and suggesting that this association may mediate the localization of sigma NS to viral factories in infected cells. We have previously shown that mu NS residues 1 to 40 or 41 are both necessary and sufficient for mu NS association with the viral microtubule-associated protein mu 2. In the present study we found that this same region of micro NS is required for its association with sigma NS. We further dissected this region, identifying residues 1 to 13 of mu NS as necessary for association with sigma NS, but not with mu 2. Deletion of sigma NS residues 1 to 11, which we have previously shown to be required for RNA binding by that protein, resulted in diminished association of sigma NS with mu NS. Furthermore, when treated with RNase, a large portion of sigma NS was released from mu NS coimmunoprecipitates, suggesting that RNA contributes to their association. The results of this study provide further evidence that mu NS plays a key role in forming the reovirus factories and recruiting other components to them.


Protective Effect of Astragaloside IV on Hepatic Injury Induced by Iron Overload.
Suitable content of iron is essential for human body, but iron overload is associated with many kinds of diseases including chronic liver damage. Recently, researchers find that iron overload promotes hepatocyte autophagy and apoptosis. However, the mechanism of iron overload in liver damage remains unclear. In this study, Lo2 cells were selected as the research object, iron dextran was a model drug, and astragaloside IV was a therapeutic drug to explore the role of iron overload. MTT assay and Annexin/PI double staining were used to measure cell viability and apoptosis. Ultrastructure was observed by transmission electron microscopy. The expression levels of apoptosis and autophagy-related proteins were determined by real-time PCR and Western Blot. The results showed that iron dextran could significantly inhibit Lo2 cell viability and increase the apoptosis rate, while astragaloside IV could reverse the inhibition of Lo2 cell viability and decrease the apoptosis rate. Transmission electron microscopy showed a significant increase in the number of autophagosomes after administration of iron dextran, and the application of astragaloside IV reduced the production of autophagosomes. LC3II/I was significantly upregulated in the model group but decreased in the astragaloside IV treatment group, and P62 showed the opposite trend. Iron dextran significantly upregulated the expression of Bax and downregulated Bcl2, while astragaloside IV reversed this trend. Finally, the inhibition of hepcidin caused by iron dextran was counteracted by astragaloside IV. In conclusion, the experimental results show that the iron overload model mainly induces excessive autophagy and apoptosis of hepatocytes, thus causing damage to hepatocytes, but astragaloside IV plays a certain therapeutic role in reversing this damage.


Mechanistic links between systemic hypertension and open angle glaucoma.
Systemic hypertension or hypertension is a very common chronic age-related disease worldwide. It is typically characterised by a sustained elevation of blood pressure, particularly when the systolic blood pressure and/or diastolic blood pressure are of more than 140 mmHg and 90 mmHg, respectively. If hypertension is not well controlled, it may lead to an increased risk of stroke and heart attack. It has been shown that hypertension is linked to various ocular diseases, including cataract, diabetic retinopathy, age-related macular degeneration, and glaucoma. Glaucoma is the leading cause of irreversible blindness worldwide. Primary open angle glaucoma is the most common form of the disease and is usually characterised by an increase in intraocular pressure. This condition, together with normal tension glaucoma, constitutes open angle glaucoma. Systemic hypertension has been identified as a risk factor for open angle glaucoma. It is speculated that blood pressure is involved in the pathogenesis of open angle glaucoma by altering intraocular pressure or ocular blood flow, or both. Recent evidence has shown that both extremely high and low blood pressure are associated with increased risk of open angle glaucoma. Additional pathogenic mechanisms, including increased inflammation likely to be involved in the development and progression of these two diseases, are discussed.


Efficacy and safety analysis of trastuzumab and paclitaxel based regimen plus carboplatin or epirubicin as neoadjuvant therapy for clinical stage II-III, HER2-positive breast cancer patients: a phase 2, open-label, multicenter, randomized trial.
This trial was designed to compare the efficacy and safety between epirubicin (E) and carboplatin (C) in combination with paclitaxel (P) and trastuzumab (H) in neoadjuvant setting. In 13 Chinese cancer centers, 100 patients with HER2-positive, locally advanced breast cancer were 1:1 randomized to receive medication as follows: trastuzumab and paclitaxel weekly combined with carboplatin weekly for PCH group, or epirubicin every 3 weeks for PEH group. Patients were given 4 to 6 cycles of chemotherapy. The primary endpoint was pathologic complete response (pCR) rate, which was no significant difference in PCH and PEH regimen (39.1% vs. 48.8%; p=0.365). However, PEH regimen achieved higher pCR in luminal-B (HER2-poitive) subgroup (55.0% vs. 24.0%; p = 0.033), but not in ERBB2+ subgroup (42.9% vs. 57.1%; p = 0.355). PEH regimen showed a favorable efficacy in PIK3CA mutated subgroup (69.2% vs.23.5%, p=0.012). No significant difference was observed in the subgroup analysis of TP53 mutation status, PTEN expression, FCGR2A SNP and FCGR3A SNP. Both regimens as neoadjuvant chemotherapy achieve similar efficacy and safety. PEH might improve pCR rate, especially in the luminal-B subtype and PIK3CA mutation subtype. PEH is feasible and less likely to increase the incidence of acute cardiac events compared to PCH.


Treatment of advanced non-small-cell lung cancer in the elderly: results of an international expert panel.
The best treatment for elderly patients with advanced non-small-cell lung cancer (NSCLC) is still debated. To guide clinical management of these patients and suggest the priorities for clinical research in this field, an International Expert Panel met in Naples, Italy, on April 19 to 20, 2004. Results and conclusions based on a review of evidence available in the literature to date are presented in this article. A comprehensive geriatric assessment is recommended to better define prognosis and to predict tolerance to treatment. In the first randomized study dedicated to elderly NSCLC patients, single-agent vinorelbine showed superiority over supportive care alone, both in terms of survival and quality of life. In a large randomized trial, gemcitabine plus vinorelbine failed to show any advantage over either agent alone. Subset analyses suggest that the efficacy of platinum-based combination chemotherapy is similar in fit older and younger patients, with an acceptable increase in toxicity for elderly patients. These data should be interpreted cautiously because retrospective subgroup analyses are encumbered by selection bias; hence, randomized trials dedicated to platinum-based chemotherapy for nonselected elderly patients are warranted. Several promising biologic therapies are under investigation; however, with present data, target-based agents as first-line treatment for elderly NSCLC patients are not yet recommended. Clinical research, with trials specifically designed for elderly patients, is mandatory. With the current evidence, single-agent chemotherapy with a third-generation drug (vinorelbine, gemcitabine, a taxane) should be the recommended option for nonselected elderly patients with advanced NSCLC. Platinum-based chemotherapy is a viable option for fit patients with adequate organ function. Best supportive care remains important, in addition to chemotherapy or as the exclusive option for patients who are unsuitable for more aggressive treatment.


[Semi-closed endarterectomy from the aortic-iliac-femoral segment].
Presented herein is a review of the literature dedicated to extended semi-closed endarterectomy from the aortoiliac segment. Considered in the article are historical data concerning the development of semi-closed loop endarterectomy, also showing present-day approaches to revascularization of the arteries of the aortofemoral zone. Also given are indications/or and contraindications to surgical management and the data on immediate and remote outcomes of semi-closed endarterectomy, which are not inferior to the classical aortofemoral reconstructions. The advantages of this method are as follows: a minimally traumatic nature, a possibility to be performed in patients presenting with severe concomitant pathology, and no use of synthetic materials.


Specific adaptation.
In this paper the key to the diagnosis, treatment and prophylaxis of chronic illnesses based on individual susceptibility to foods, drugs, food-drug combinations and environmental chemicals is discussed thoroughly.


Rituximab therapy leads to rapid decline of serum IgG4 levels and prompt clinical improvement in IgG4-related systemic disease.
Patients with IgG4-related systemic disease (IgG4-RSD) frequently show an incomplete response to treatment with glucocorticoids and traditional disease-modifying antirheumatic drugs (DMARDs). B lymphocyte depletion is a therapeutic strategy known to be effective for pemphigus vulgaris, an autoimmune condition mediated by IgG4 autoantibodies. This study was performed to assess the clinical and serologic responses to B lymphocyte depletion therapy with rituximab in patients with IgG4-RSD. Four patients with IgG4-RSD were treated with 2 intravenous doses (1 gram each) of rituximab. Clinical improvement was assessed by monitoring the tapering/discontinuation of prednisone and DMARDs, and by measuring the serum concentrations of B lymphocytes, immunoglobulins, and IgG subclasses before and after therapy. Clinical features of IgG4-RSD in these 4 patients included autoimmune pancreatitis, sclerosing cholangitis, lymphoplasmacytic aortitis, salivary gland involvement, orbital pseudotumor, and lacrimal gland enlargement. The 3 patients with elevated serum IgG and IgG4 levels at baseline had a mean IgG concentration of 2,003 mg/dl (normal range 600-1,500 mg/dl) and a mean IgG4 concentration of 2,160 mg/dl (normal range 8-140 mg/dl). Among these patients, the serum IgG4 concentrations declined by a mean of 65% within 2 months of rituximab administration. All 4 patients demonstrated striking clinical improvement within 1 month of the initiation of rituximab therapy, and tapering or discontinuation of their treatment with prednisone and DMARDs was achieved in all 4 patients. A decrease in IgG concentration was observed for the IgG4 subclass only. Treatment with rituximab led to prompt clinical and serologic improvement in these patients with refractory IgG4-RSD, and is a viable treatment option for this condition. The decline in serum IgG4 concentrations was substantially steeper than that of the autoantibody concentrations in immune-mediated conditions in which rituximab is effective, such as in rheumatoid arthritis. In addition, the reduction in IgG-subclass levels appeared to be specific for IgG4. The swift improvement of IgG4-RSD suggests that rituximab achieves its effects in IgG4-RSD by depleting the pool of B lymphocytes that replenish short-lived IgG4-secreting plasma cells.


Dermal, oral, and inhalation pharmacokinetics of methyl tertiary butyl ether (MTBE) in human volunteers.
Methyl tertiary butyl ether (MTBE), a gasoline additive used to increase octane and reduce carbon monoxide emissions and ozone precursors, has contaminated drinking water and can lead to exposure by oral, inhalation, and dermal routes. To determine its dermal, oral, and inhalation kinetics, 14 volunteers were exposed to 51.3 microg/ml MTBE dermally in tap water for 1 h, drank 2.8 mg MTBE in 250 ml Gatorade(R), and inhaled 3.1 ppm. MTBE for 1 h. Blood and exhaled breath samples were then obtained. Blood MTBE peaked between 15 and 30 min following oral exposure, at the end of inhalation exposure, and ~5 min after dermal exposure. Elimination by each route was described well by a three-compartment model (Rsq >0.9). The Akaike Information Criterion for the three-compartment model was smaller than the two-compartment model, supporting it over the two-compartment model. One metabolite, tertiary butyl alcohol (TBA), measured in blood slowly increased and plateaued, but it did not return to the pre-exposure baseline at the 24-h follow-up. TBA is very water-soluble and has a blood:air partition ratio of 462, reducing elimination by exhalation. Oral exposure resulted in a significantly greater MTBE metabolism into TBA than by other routes based on a greater blood TBA:MTBE AUC ratio, implying significant first-pass metabolism. The slower TBA elimination may make it a better biomarker of MTBE exposure, though one must consider the exposure route when estimating MTBE exposure from TBA because of first-pass metabolism. Most subjects had a baseline blood TBA of 1-3 ppb. Because TBA is found in consumer products and can be used as a fuel additive, it is not a definitive marker of MTBE exposure. These data provide the risk assessment process of pharmacokinetic information relevant to the media through which most exposures occur-air and drinking water.


The Effects of Secondary Cleft Procedures on Alar Base Position and Nostril Morphology in Patients with Unilateral Clefts.
To compare effects of secondary cleft procedures on alar base position and nostril morphology. Retrospective review. Multidisciplinary cleft clinic at tertiary center. Seventy consecutive patients with unilateral clefts were grouped according to secondary procedure. Alveolar bone graft versus total lip takedown with anatomic muscle repair versus single-stage total lip with cleft septorhinoplasty (nose-lip) versus rhinoplasty alone. Anthropometric measurements were recorded from pre- and postoperative photographs. Ratios of cleft to noncleft side were compared within and across groups pre- and postoperatively using parametric and nonparametric tests. Within the bone graft group, no differences were seen postoperatively in alar base position in long-term follow-up. The total lip group demonstrated greater symmetry at the alar base (P < .001), increased vertical lip dimension (P < .001), and decreased nostril height (P = .004) postoperatively. Within the nose-lip group, increased vertical dimension and alar base support (P < .001) were also seen postoperatively. Across groups, the single-stage nose-lip group demonstrated greatest alar base symmetry on worm's-eye view (P < .04). Alar base asymmetry in patients with unilateral clefts may be related to soft tissue deficiency and was not affected by alveolar bone grafting. Total lip takedown with anatomic muscle reapproximation was associated with increased alar base symmetry and vertical lip dimension on cleft to noncleft side. Greatest symmetry at the alar base was seen following single-stage nose-lip reconstruction, which may be an effective technique for correcting the secondary cleft lip nasal deformity.


Chromatographic pattern classification.
In this paper, we propose and evaluate methodologies for the classification of images from thin-layer chromatography. Each individual sample is characterized by an intensity profile that is further represented into a feature space. The first steps of this process aim at obtaining a robust estimate of the intensity profile by filtering noise, reducing the influence of background changes, and by fitting a mixture of Gaussians. The resulting profiles are represented by a set of appropriate features trying to characterize the state of nature, here spread out over four classes, one for normal subjects and the other three corresponding to lysosomal diseases, which are disorders responsible for severe nerve degeneration. For classification purposes, a novel solution based on a hierarchical structure is proposed. The main conclusion of this paper is that an automatically generated decision tree presents better results than more conventional solutions, being able to deal with the natural imbalance of the data that, as consequence of the rarity of lysosomal disorders, has very few representative cases in the disease classes when compared with the normal population.


Cost-effectiveness analysis of a new 8% capsaicin patch compared to existing therapies for postherpetic neuralgia.
The purpose of this study was to compare the cost effectiveness of a new 8% capsaicin patch, compared to the current treatments for postherpetic neuralgia (PHN), including tricyclic antidepressants (TCAs), topical lidocaine patches, duloxetine, gabapentin, and pregabalin. A 1-year Markov model was constructed for PHN with monthly cycles, including dose titration and management of adverse events. The perspective of the analysis was from a payer perspective, managed-care organization. Clinical trials were used to determine the proportion of patients achieving at least a 30% improvement in PHN pain, the efficacy parameter. The outcome was cost per quality-adjusted life-year (QALY); second-order probabilistic sensitivity analyses were conducted. The effectiveness results indicated that 8% capsaicin patch and topical lidocaine patch were significantly more effective than the oral PHN products. TCAs were least costly and significantly less costly than duloxetine, pregabalin, topical lidocaine patch, 8% capsaicin patch, but not gabapentin. The incremental cost-effectiveness ratio for the 8% capsaicin patch overlapped with the topical lidocaine patch and was within the accepted threshold of cost per QALY gained compared to TCAs, duloxetine, gabapentin, and pregablin. The frequency of the 8% capsaicin patch retreatment assumption significantly impacts its cost-effectiveness results. There are several limitations to this analysis. Since no head-to-head studies were identified, this model used inputs from multiple clinical trials. Also, a last observation carried forward process was assumed to have continued for the duration of the model. Additionally, the trials with duloxetine may have over-predicted its efficacy in PHN. Although a 30% improvement in pain is often an endpoint in clinical trials, some patients may require greater or less improvement in pain to be considered a clinical success. The effectiveness results demonstrated that 8% capsaicin and topical lidocaine patches had significantly higher effectiveness rates than the oral agents used to treat PHN. In addition, this cost-effectiveness analysis found that the 8% capsaicin patch was similar to topical lidocaine patch and within an accepted cost per QALY gained threshold compared to the oral products.


Effects of inorganic selenium injection on the performance of beef cows and their subsequent calves.
Sixty-seven pregnant Angus cows and their subsequent calves were used in a randomized design to evaluate the effect of inorganic Se injection from 80 d of gestation until weaning on cow reproductive parameters, BW and BCS evolution, milk yield and quality, calf growth and calf hematology parameters. The treatments were as follows: 1) Se+: cows were administered Se as sodium selenite at doses of 0.05 mg/Kg of BW at 80, 140, 200, 260 d of gestation and 30, 90, 150 and 240 d of lactation. Calves from Se + cows were Se as sodium selenite subcutaneously injected with a dose of 0.05 mg/Kg of BW at 70, 104, 144 d of age; 2) Control: cows and calves were administered sterile NaCl solution (9 g/l) at the same volume and intervals as Se + treatment. At the start of the experiment, forage Se concentration was 58.6 ppb ± 7.6 ppb, and cow whole blood Se concentration was similar (P > 0.10) between treatments (Se+, 29.3 ± 0.3 ppb; Control, 28.1 ± 0.5 ppb). After Se injection, cow whole blood Se concentration was increased (P < 0.01) in Se + cows in relation to control cows until the end of the experiment. Calf whole blood Se concentration was increased (P < 0.01) at birth and 30 d of age in calves from Se + cows compared to calves from Control cows. However, calf whole blood Se concentration was not different (P > 0.10) between treatments at d 70 after birth. After the first Se injection (70 d of age), calf whole blood Se concentration was increased (P < 0.01) at 104 and 144 d of age in calves from Se + cows compared to calves from Control cows. Cow BW and BCS did not differ (P > 0.10) between treatments throughout the experiment. Ovarian follicle diameter measured by ultrasound 47 d postpartum was greater (P = 0.03) in Se + cows compared to Control cows. Pregnancy rate to fixed time artificial insemination (FTAI), overall pregnancy rate (P > 0.10) and pregnancy loss (P = 0.19) did not differ between groups. Selenium injection did not affect (P > 0.10) milk yield and composition. Calf morphometric parameters, BW at birth and growth rate during lactation were not affected (P > 0.10) by treatments. No significant effect (P > 0.10) was observed between treatments on hematological results of blood samples from calves at weaning. Results of this study shown that inorganic Se injection was an efficient tool to improve Se-status in cow-calf operation under extensive management. Selenium injection increased follicle diameter in postpartum beef cow, however, did not affect the rest of the variables studied.


Factors associated with insomnia and complementary medicine use in children: results of a national survey.
Sleep difficulties are a serious health problem in children, and interest in using complementary and alternative medicine (CAM) therapies to treat sleep is growing. In this study, we aimed to identify: the prevalence of sleep difficulties in children, and the prevalence and patterns of CAM use among children with trouble sleeping. We used the 2012 National Health Interview Survey (NHIS) dataset to estimate the prevalence of sleep difficulties and CAM use in children ages 6-17 years. Prevalence estimates were weighted to reflect the survey's sampling design. We used logistic regression to explore associations between sleep difficulties, psychosocial factors, comorbidities and CAM use. 6.4% of children in the 2012 NHIS dataset reported regular difficulty sleeping in the last year, corresponding to an estimated 1.5 million children in the US. Older age, poorer health status, more missed school days, and multiple comorbidities were all associated with sleep difficulties (p ≤ 0.001). Among children with sleep difficulties, 29% used at least one CAM therapy. Of the CAM therapies surveyed, non-vitamin, non-mineral supplements were the most commonly used (14.6%), followed by manipulation therapies (9.2%) and mind-body techniques (8.8%). Parental education and CAM use were most strongly associated with child CAM use (p ≤ 0.001). CAM therapies, particularly non-vitamin, non-mineral supplements, are commonly used among children with sleeping problems. More research is needed to characterize the safety and efficacy of CAM therapies for sleep in this population.


Analysis of receptor tyrosine kinase ROS1-positive tumors in non-small cell lung cancer: identification of a FIG-ROS1 fusion.
To deepen our understanding of mutant ROS1 expression, localization, and frequency in non-small cell lung cancer (NSCLC), we developed a highly specific and sensitive immunohistochemistry (IHC)-based assay that is useful for the detection of wild-type and mutant ROS1. We analyzed 556 tumors with the ROS1 D4D6 rabbit monoclonal antibody IHC assay to assess ROS1 expression levels and localization. A subset of tumors was analyzed by FISH to determine the percentage of these tumors harboring ROS1 translocations. Using specific and sensitive IHC assays, we analyzed the expression of anaplastic lymphoma kinase (ALK), EGFR L858R, and EGFR E746-A750del mutations in a subset of lung tumors, including those expressing ROS1. In our NSCLC cohort of Chinese patients, we identified 9 (1.6%) tumors expressing ROS1 and 22 (4.0%) tumors expressing ALK. FISH identified tumors with ALK or ROS1 rearrangements, and IHC alone was capable of detecting all cases with ALK and ROS1 rearrangements. ROS1 fusion partners were determined by reverse transcriptase PCR identifying CD74-ROS1, SLC34A2-ROS1, and FIG-ROS1 fusions. Some of the ALK and ROS1 rearranged tumors may also harbor coexisting EGFR mutations. NSCLC tumors with ROS1 rearrangements are uncommon in the Chinese population and represent a distinct entity of carcinomas. The ROS1 IHC assay described here is a valuable tool for identifying patients expressing mutant ROS1 and could be routinely applied in clinical practice to detect lung cancers that may be responsive to targeted therapies.


Feasibility of surface-enhanced Raman spectroscopy for rapid detection of aflatoxins in maize.
Rapid and sensitive surface-enhanced Raman spectroscopy (SERS) for aflatoxin detection was employed for development of the models to classify and quantify aflatoxin levels in maize at concentrations of 0 to 1,206 μg/kg. Highly effective SERS substrate (Ag nanosphere) was prepared and mixed with a sample extract for SERS measurement. Strong Raman bands associated with aflatoxins and changes in maize kernels induced by aflatoxin contamination were observed in different SERS spectroscopic regions. The k-nearest neighbors (KNN) classification model yielded high classification accuracy and lower prediction error with no misclassification of contaminated samples as aflatoxin negative. The multiple linear regression (MLR) models showed a higher predictive accuracy with stronger correlation coefficients (r = 0.939-0.967) and a higher sensitivity with lower limits of detection (13-36 μg/kg) and quantitation (44-121 μg/kg) over other quantification models. Paired sample t test exhibited no statistically significant difference between the reference values and the predicted values of SERS in most chemometric models. The proposed SERS method would be a more effective and efficient analytical tool with a higher accuracy and lower constraints for aflatoxin analysis in maize compared to other existing spectroscopic methods and conventional Raman spectroscopy.


The state of the art. Donor site harvest, graft yield estimation, and recipient site preparation for follicular-unit hair transplantation.
The field of hair transplantation has moved beyond the provision of a dense frontal hairline to techniques that are designed to provide a natural appearance in as few sessions as possible. The concept of the follicular unit has revolutionized the preparation of both the grafts and the recipient sites. We will describe the mathematics of hair restoration and our techniques for determining the length and height of the donor strip for a given number of desired follicular-unit grafts. We will also elucidate our technique for cutting the follicular-unit grafts using microscopes and transillumination. We now use specially designed blades or punches to make shallow slits or slots for recipient sites. Striving for teamwork and efficiency is of paramount importance in hair transplantation, and we will describe our coordination and training methods as well as the role of the surgeon.


Shared sequence characteristics identified in non-canonical rearrangements of HSV-1 genomes.
Mutations and genetic rearrangements are the primary driving forces of evolution. Viruses provide valuable model systems for investigating these mechanisms due to their rapid evolutionary rates and vast genetic variability. To investigate genetic rearrangements in the double-stranded DNA genome of herpes simplex virus type 1, the viral population was serially passaged in various cell types. The serial passaging led to formation of defective genomes, resulted from cell-specific non-canonical rearrangements (NCRs). Interestingly, we discovered shared sequence characteristics underlying the formation of these NCRs across all cell types. Moreover, most NCRs identified in clinical samples shared these characteristics. Based on our findings, we propose a model elucidating the formation of NCRs during viral replication within the nucleus of eukaryotic cells.


A novel HLA (HLA-A*0201) transgenic rabbit model for preclinical evaluation of human CD8+ T cell epitope-based vaccines against ocular herpes.
We introduced a novel humanized HLA-A*0201 transgenic (HLA Tg) rabbit model to assess the protective efficacy of a human CD8(+) T cell epitope-based vaccine against primary ocular herpes infection and disease. Each of the three immunodominant human CD8(+) T cell peptide epitopes from HSV-1 glycoprotein D (gD(53-61), gD(70-78), and gD(278-286)) were joined with a promiscuous human CD4(+) T cell peptide epitope (gD(49-82)) to construct three separate pairs of CD4-CD8 peptides. Each CD4-CD8 peptide pair was then covalently linked to an N(epsilon)-palmitoyl-lysine residue via a functional base lysine amino group to construct CD4-CD8 lipopeptides. HLA Tg rabbits were immunized s.c. with a mixture of the three CD4-CD8 HSV-1 gD lipopeptides. The HSV-gD-specific T cell responses induced by the mixture of CD4-CD8 lipopeptide vaccine and the protective efficacy against acute virus replication and ocular disease were determined. Immunization induced HSV-gD(49-82)-specific CD4(+) T cells in draining lymph node (DLN); induced HLA-restricted HSV-gD(53-61), gD(70-78), and gD(278-286)-specific CD8(+) T cells in DLN, conjunctiva, and trigeminal ganglia and reduced HSV-1 replication in tears and corneal eye disease after ocular HSV-1 challenge. In addition, the HSV-1 epitope-specific CD8(+) T cells induced in DLNs, conjunctiva, and the trigeminal ganglia were inversely proportional with corneal disease. The humanized HLA Tg rabbits appeared to be a useful preclinical animal model for investigating the immunogenicity and protective efficacy of human CD8(+) T cell epitope-based prophylactic vaccines against ocular herpes. The relevance of HLA Tg rabbits for future investigation of human CD4-CD8 epitope-based therapeutic vaccines against recurrent HSV-1 is discussed.


[Pleomorphic carcinoma of the lung heralded as bilateral leg pain].
This is a case of a 70 year old male patient suffering from bilateral leg pain for 2 months. Physical examination disclosed clubbing. X-rays of the legs showed bilateral periosteal elevation with subperiosteal bone formation. 99TM-diphosphonate bone scan was negative. A search for malignancy revealed pleomorphic carcinoma of the right lung. Pain symptom disappeared 2 days after resection of the tumor. Repeated X-rays of the legs, three and a half months later showed no change and the clubbing persisted. Hence, unexplained bilateral leg pain should raise suspicion of hypertrophic osteoarthropathy, and elicit a search for secondary disease especially lung tumor. It is interesting to point out the negative bone scan and the rapid resolution of patient symptoms after resection of the tumor.


Adjuvant effect of IL-12: conversion of peptide antigen administration from tolerizing to immunizing for CD8+ T cells in vivo.
CD8+ T cells from TCR transgenic 2C mice, specific for SIYRYYGL peptide bound to H-2Kb, were adoptively transferred into C57BL/6 recipients to allow monitoring of their location, numbers, and phenotype upon peptide challenge. Recipients were primed by s.c. injection of SIYRYYGL alone or with CFA or IL-12, and the transferred cells then tracked by flow cytometry using the 1B2 mAb specific for the 2C TCR. Peptide alone induced a transient and weak expansion of 1B2+ cells in the draining lymph nodes (DLN) by day 3, but these cells were tolerant to secondary peptide challenge. In contrast, priming with CFA/peptide resulted in a large clonal expansion of 1B2+ cells in DLN by day 3, and the cells exhibited a CD25highCD44high phenotype, blast transformation, and lytic effector function. By day 5, 1B2+ cell numbers decreased in the DLN and increased in the spleen and blood. 1B2+ cells with a memory phenotype persisted through day 60 in the DLN, spleen, and blood and responded to secondary peptide challenge. Immunization with peptide, along with IL-12, mimicked the adjuvant effects of CFA with respect to phenotype, clonal expansion, effector function, and establishment of memory. IL-12 was not unique in providing this adjuvant effect however, since CFA/peptide immunization of IL-12-deficient recipient mice also resulted in 1B2+ T cell activation and clonal expansion. Thus, CFA or IL-12 can enhance Ag-specific CD8+ T cell responses to peptide, demonstrating that an inflammatory cytokine(s) can support activation and prevent tolerance induction.


Tissue Remodeling in Ocular Mucous Membrane Pemphigoid.
Ocular mucous membrane pemphigoid (OcMMP) is a rare eye disease characterized by relapsing-remitting or persisting long-lasting inflammatory events associated with progressive scarring. Despite long-term immunomodulating therapy, abnormal fibrosis keeps worsening in patients with OcMMP. This study investigates the fibrotic process in patients with OcMMP, as well as the critical role of the epithelium in modulating the local fibrosis. In this prospective, observational pilot study, patients affected by long-lasting OcMMP were compared with age- and gender-matched healthy controls. Clinical grading was assessed, and conjunctival biopsy and impression cytology were performed. Conjunctival samples were used for quantifying the expression of transcripts regulating the inflammatory and fibrogenic processes. Ocular surface clinical and functional markers worsened in patients with OcMMP with fibrotic disease progression. In more advanced disease stages, both impression cytologies and conjunctival biopsies revealed increased tissue remodeling and profibrotic markers (α-SMA and TGF-β), and decreased levels of inflammatory markers (I-CAM1, IL-10, and IL-17). Increased epithelial expression of profibrotic markers and histological changes were detected. Chronic OcMMP is characterized by a progressive, aberrant self-sustaining fibrotic process that worsens clinical signs and symptoms. Conjunctival epithelial cells may transdifferentiate into myofibroblast-like phenotypes when chronically exposed to high levels of inflammation, as in the case of OcMMP. Tissue remodeling markers in OcMMP could be used as early diagnostic, prognostic, and therapeutic biomarkers, harvested in a non-invasive and painless procedure such as impression cytologies.