Structural Identification of Monounsaturated Branched Chain Fatty Acid Methyl Esters by Combination of Electron Ionization and Covalent Adduct Chemical Ionization Tandem Mass Spectrometry.
Monounsaturated normal fatty acids (n-MUFA) and saturated branched chain fatty acids (BCFA) are structurally characterized by separate tandem mass spectrometry methods for double bond localization and for chain branching in their respective fatty acid methyl ester (FAME) derivatives; however, these methods have never been applied to branched monounsaturated FAME. Here, we report application of electron ionization (EI)-MS/MS and solvent-mediated covalent adduct chemical ionization (CACI)-MS/MS of monounsaturated BCFA methyl esters (MUBCFAME) of a chain length of 15-20 carbons. A novel system was used to implement CI with low vapor pressure reagents in a tabletop triple quadrupole MS. Anteiso-MUBCFA EI-MS/MS of the molecular ion (M) yields a characteristic diagnostic ion [M-29]+. iso-MUBCFA can be distinguished from n-MUFA by an ion intensity ratio of [M-32]+/[M-43]+, with iso-MUBCFA yielding a ratio greater than 1.7, while n-MUFA yields a ratio less than 1.0. Chain branching at the iso and anteiso positions, terminal isopropyl and sec-butyl, respectively, do not alter CACI-MS/MS diagnostic ions compared to normal BCFA, enabling double bond positions of MUBCFA to be determined with the analogous α and ω diagnostic ions from cleavage on both sides of the erstwhile double bond. Taken together, this straightforward FAME-based technique via combination of EI-MS/MS and CACI-MS/MS enables fundamental structural identification of MUBCFA without standards.


Photodynamic therapy with phthalocyanine sensitisation: quantitative studies in a transplantable rat fibrosarcoma.
Photodynamic therapy (PDT) is a promising approach to the local destruction of malignant tumours, but little work has been done to determine which factors control the extent of tissue necrosis produced. Using a new photosensitiser, a sulphonated aluminium phthalocyanine (AlSPc) and light from an argon ion pumped dye laser at 675 nm, we quantified the effects of interstitial PDT in a transplantable fibrosarcoma in rats. At 100mW laser power, thermal effects were comparable to those of PDT, so subsequent studies were carried out at 50 mW, where thermal effects were minimal. The depth of PDT necrosis increased with the logarithm of the applied energy. Tissue concentration of AlSPc was measured by alkali extraction and at all times after sensitisation, correlated well with the necrosis produced with a given light dose. Peak tumour concentration of AlSPc occurred 24-48 h after sensitisation compared with a peak at 3 h in muscle. The peak ratio tumour:muscle was 2:1 at 24 h. Apart from a different time interval to reach the peak sensitiser concentration, the extent of tumour damage varied with the light and sensitiser parameters in a similar way to that found in normal liver, although the optical penetration depth was greater in the tumour (2.5 mm vs. 1.8 mm). At doses of AlSPc below 1 mg kg-1 the diameter of necrosis increased with the logarithm of the dose of sensitiser, and doubling the dose from 0.25 to 0.5 mg kg-1 increased the depth of necrosis by 50%. However, at higher doses, the changes were smaller and increasing the dose from 2.5 to 5 mg kg-1 only increased the necrosis by 10% for the same light dose. In all dose ranges, a given percentage increase in the tissue concentration of AlSPc gave a much smaller percentage increase in the extent of necrosis for the same light dose, suggesting that selectivity of necrosis between tumour and normal tissue is likely to be much less than the selectivity of retention of the photosensitiser. From these results, the extent of PDT necrosis in this fibrosarcoma is as predictable as it is in normal liver if the light dose, tissue concentration of AlSPc and optical penetration depth of the tissue are known. Further studies are now required on different tumour models to establish how tumours respond compared with adjacent normal tissue when the tumour is growing in its organ of origin rather than the non-physiological situation using a transplantable tumour as in this study.


A library of oligosaccharide probes (neoglycolipids) from N-glycosylated proteins reveals that conglutinin binds to certain complex-type as well as high mannose-type oligosaccharide chains.
This report describes the preparation of a library of oligosaccharide probes (neoglycolipids) from N-glycosylated proteins, characterization of the probes by liquid secondary ion mass spectrometry, and investigation of their reactions with 125I-labeled bovine serum conglutinin by chromatogram binding assays. The results, together with additional binding studies using neoglycolipids derived from purified complex type bi-, tri-, and tetraantennary oligosaccharides from urine, or their glycosidase-treated products, have shown that the combining specificity of conglutinin includes structures not only on high mannose-type oligosaccharides but also on hybrid- and complex-type chains. With high mannose-type oligosaccharides there is increased reactivity from the Man5 to the Man8 structures, indicating a preference for the terminal Man alpha 1-2 sequence. With complex- and hybrid-type oligosaccharides, the requirements for binding are the presence of nonreducing terminal N-acetylglucosamine or mannose residues, but the presence of a bisecting N-acetylglucosamine residue may inhibit binding. From these results it is deduced that the reactivity of conglutinin with the complement glycopeptide iC3b rather than the intact glycoprotein C3 is due to the oligosaccharide accessibility rendered by proteolysis in the complement cascade.


Accidental Mycobacterium bovis infection in a veterinarian.
A veterinarian developed tenosynovitis and secondary carpal tunnel syndrome following accidental inoculation of Mycobacterium bovis during the necropsy of a tuberculous possum from Westland, New Zealand. M. bovis infection is a zoonotic disease, and occupational exposure to tuberculous animals places people at risk of contracting the disease. Adhering to safe work practices reviewed in this article is important to minimise the risk of infection to people handling tuberculous animals.


Limited proteolysis of native proteins: the interaction between avidin and proteinase K.
Avidin is a tetramer of 16-kDa subunits that have a high affinity for biotin. Proteolysis of native apoavidin by proteinase K results in a limited attack at the loop between beta-strands 3 and 4, involving amino acids 38-43. Specifically, sites of proteolysis are at Thr 40-Ser 41 and Asn 42-Glu 43. The limited proteolysis results in an avidin product that remains otherwise intact and which has enhanced binding for 4'-hydroxyazobenzene-2-benzoic acid (HABA), a chromogenic reporter that can occupy the biotin-binding site. Saturation of the biotin-binding site with the natural ligand protects avidin from proteolysis, but saturation with HABA enhances the rate of proteolysis of the same site. Analysis of the three-dimensional structures of apoavidin and holoavidin reveals that the 3-4 loop is accessible to solvent and scores highly in an algorithm developed to identify sites of proteolytic attack. The structure of holoavidin is almost identical to the apoprotein. In particular, the 3-4 loop has the same structure in the apo and holo forms, yet there are marked differences in proteolytic susceptibility of this region. Evidence suggests that the 3-4 loop is rather mobile and flexible in the apoprotein, and that it becomes constrained upon ligand binding. In one crystal structure of the apoprotein, this loop appears constrained by contacts with symmetry-related molecules. Structural analyses suggest that the "lid" to the biotin-binding site, formed by the 3-4 loop, is displaced and made more accessible by HABA binding, thereby enhancing its proteolytic susceptibility.


Fronto-striatal dopamine D2 receptor availability is associated with cognitive variability in older individuals with low dopamine integrity.
Within-person, moment-to-moment, variability in behavior increases with advancing adult age, potentially reflecting the influence of reduced structural and neurochemical brain integrity, especially that of the dopaminergic system. We examined the role of dopamine D2 receptor (D2DR) availability, grey-, and white-matter integrity, for between-person differences in cognitive variability in a large sample of healthy older adults (n = 181; 64-68 years) from the Cognition, Brain, and Aging (COBRA) study. Intra-individual variability (IIV) in cognition was measured as across-trial variability in participants' response times for tasks assessing perceptual speed and working memory, as well as for a control task of motor speed. Across the whole sample, no associations of D2DR availability, or grey- and white-matter integrity, to IIV were observed. However, within-person variability in cognition was increased in two subgroups of individuals displaying low mean-level cognitive performance, one of which was characterized by low subcortical and cortical D2DR availability. In this latter group, fronto-striatal D2DR availability correlated negatively with within-person variability in cognition. This finding suggests that the influence of D2DR availability on cognitive variability may be more easily disclosed among individuals with low dopamine-system integrity, highlighting the benefits of large-scale studies for delineating heterogeneity in brain-behavior associations in older age.


Mutant ataxin-3 with an abnormally expanded polyglutamine chain disrupts dendritic development and metabotropic glutamate receptor signaling in mouse cerebellar Purkinje cells.
Spinocerebellar ataxia type 3 (SCA3) is caused by the abnormal expansion of CAG repeats within the ataxin-3 gene. Previously, we generated transgenic mice (SCA3 mice) that express a truncated form of ataxin-3 containing abnormally expanded CAG repeats specifically in cerebellar Purkinje cells (PCs). Here, we further characterize these SCA3 mice. Whole-cell patch-clamp analysis of PCs from advanced-stage SCA3 mice revealed a significant decrease in membrane capacitance due to poor dendritic arborization and the complete absence of metabotropic glutamate receptor subtype1 (mGluR1)-mediated retrograde suppression of synaptic transmission at parallel fiber terminals, with an overall preservation of AMPA receptor-mediated fast synaptic transmission. Because these cerebellar phenotypes are reminiscent of retinoic acid receptor-related orphan receptor α (RORα)-defective staggerer mice, we examined the levels of RORα in the SCA3 mouse cerebellum by immunohistochemistry and found a marked reduction of RORα in the nuclei of SCA3 mouse PCs. To confirm that the defects in SCA3 mice were caused by postnatal deposition of mutant ataxin-3 in PCs, not by genome disruption via transgene insertion, we tried to reduce the accumulation of mutant ataxin-3 in developing PCs by viral vector-mediated expression of CRAG, a molecule that facilitates the degradation of stress proteins. Concomitant with the removal of mutant ataxin-3, CRAG-expressing PCs had greater numbers of differentiated dendrites compared to non-transduced PCs and exhibited retrograde suppression of synaptic transmission following mGluR1 activation. These results suggest that postnatal nuclear accumulation of mutant ataxin-3 disrupts dendritic differentiation and mGluR-signaling in SCA3 mouse PCs, and this disruption may be caused by a defect in a RORα-driven transcription pathway.


Quantitative gas chromatography-olfactometry. Analytical characteristics of a panel of judges using a simple quantitative scale as gas chromatography detector.
Some questions related to the ability of gas chromatography-olfactometry (GC-O) to provide quantitative measurements of the concentration of a given odorant in an extract are explored and discussed. A panel of eight individuals has been used to evaluate the intensity of 15 odorants present at different concentrations. The use of very simple scales, such as a 0-3 scale, makes it possible to build calibration graphs based on the different stimulus-response models (Fechner, Stevens, Hill) and, with a proper calibration, up to nine different concentration levels can be discriminated by an eight-judge panel. The signal shows a good long-term stability, and its precision varies between 3.7 and 8% of the whole scale, with 5.7% as average. Sensitivity is extremely dependent on the compound: in the best case a concentration change of 20% can be detected, while in the worst, concentrations must differ more than one order of magnitude. In average, concentrations must differ between 2 and 4.7 times (including calibration error) or between 1.2 and 2.3 (excluding calibration error). The performance of the different judges, the effect of the close elution of two odors and the benefits derived from the use of more complex scales (7-points) are briefly discussed from the perspective of the analytical performance of GC-O methods.


Removal of nuclease contamination during purification of recombinant prototype foamy virus integrase.
Retroviral infection requires integration of the viral genome into the host genome. Recombinant integrase proteins may be purified following bacterial expression. A bulk biochemical assay of integrase function relies on the conversion of supercoiled plasmids to linear or relaxed circles. Single molecule molecular tweezer assays of integrase also evaluate the conversion of supercoiled DNA to nicked and broken species. A bacterial nuclease that co-purifies with retroviral integrase may affect the quantitation of integration activity in bulk or single molecule assays. During purification of retroviral integrase from bacteria, fractions may be screened for contaminating nuclease activity. In order to efficiently separate the nuclease from integrase, the binding affinities of each protein must differ. We find that a co-purifying nuclease may be efficiently separated from integrase based on heparin affinity, but not ionic affinity.


Synthesis, biological evaluation, and docking study of indole aryl sulfonamides as aromatase inhibitors.
In order to identify new aromatase enzyme inhibitors, thirty aryl sulfonamide derivatives containing an indole nucleus have been synthesized. The enzyme inhibition assay showed that four compounds inhibit aromatase in the sub-micromolar range. Loading concentrations of these four compounds were afterwards tested for cell viability and cytotoxicity on MCF7 human breast cancer cells, revealing a time- and dose-dependent decrease of active metabolizing cells over the time of the culture (0-72 h), starting from a concentration of 100 μM. Likewise LDH released raised up to 40% at early time of exposures (24 h). Finally, the docking study showed that the best active compounds efficiently bound in the active site of the aromatase; high values of HBD and low levels of HBA are the principal requirement evidenced by the QSAR model.


T-lymphocytes in experimental autoimmune myasthenia gravis. Isolation of T-helper cell lines.
The role of T-lymphocytes in Experimental Autoimmune Myasthenia Gravis (EAMG) was investigated. We generated highly purified, acetylcholine receptor (AChR)-specific T-cell populations and subsequently characterized these cell lines with respect to their membrane phenotype and their function. Using a series of mouse monoclonal antibodies directed against rat lymphocyte surface differentiation antigens, the vast majority of line cells was shown to express a leucocyte common antigen, a T-common antigen and a T-helper antigen. Small subpopulations were Ia or T suppressor antigen-positive. Adaptive transfer to sublethally irradiated, thymectomized recipients revealed that 1 X 10(6) AChR-specific line cells could cooperate effectively with 10 X 10(6) AChR-primed, complement (C3) receptor-bearing (B-cell enriched) spleen cells in the production of anti-AChR autoantibodies. Recipients of B-cells along with relevant line cells developed an acute myasthenic syndrome 6-7 days after cell transfer. Electron-microscopical examination revealed the typical features of "acute phase" EAMG with heavy mononuclear infiltration. There was, however, no evidence antibody-independent cytotoxic activity exerted by AChR-specific line cells.


Vitamin D: A Narrative Review Examining the Evidence for Ten Beliefs.
Over the past decade, a large body of observational evidence has suggested an association between lower vitamin D status (25-hydroxyvitamin D) and multiple acute and chronic disorders, including cancer, multiple sclerosis, depression and respiratory tract infections. This evidence has fostered the hypothesis that increasing vitamin D intake may treat and prevent such disorders. Our objective was to perform a critical analysis of the highest-level evidence for ten common beliefs regarding vitamin D for the prevention of falls, fractures and respiratory tract infections, the reduction of cancer incidence/mortality and overall mortality, and the prevention or treatment of depression/mental well-being, rheumatoid arthritis and multiple sclerosis, as well as maximum dosing and regular testing. We searched the Cochrane Database of Systematic Reviews and PubMed (up to August 2014) for randomized controlled trials and systematic reviews/meta-analyses based on those studies. All searches were performed, all evidence reviewed and each section written by at least two authors. The evidence shows that vitamin D supplementation provides some benefit in fracture prevention (likely ∼10-15 % relative reduction), particularly at a dose ≥800 IU and with calcium; a likely benefit in the rate of falls, though it is less clear whether the number of fallers changes; and a possible small (∼5 %) relative reduction in mortality. Evidence does not support the use of vitamin D supplementation for the prevention of cancer, respiratory infections or rheumatoid arthritis. Similarly, evidence does not support vitamin D supplementation for the treatment of multiple sclerosis and rheumatoid arthritis or for improving depression/mental well-being. Regular testing of 25-hydroxyvitamin D is generally not required, and mega-doses (≥300,000 IU) appear to increase harms. Much of the evidence is at high risk of bias, with multiple flaws, including analyses of secondary endpoints, small and underpowered studies, inconsistent results and numerous other issues. Therefore, enthusiasm for a vitamin D panacea should be tempered.


Processing single-cell RNA-seq datasets using SingCellaR.
Single-cell RNA sequencing has led to unprecedented levels of data complexity. Although several computational platforms are available, performing data analyses for multiple datasets remains a significant challenge. Here, we provide a comprehensive analytical protocol to interrogate multiple datasets on SingCellaR, an analysis package in R. This tool can be applied to general single-cell transcriptome analyses. We demonstrate steps for data analyses and visualization using bespoke pipelines, in conjunction with existing analysis tools to study human hematopoietic stem and progenitor cells. For complete details on the use and execution of this protocol, please refer to Roy et al. (2021).


The "slide technique"-a novel free-hand method of subaxial cervical pedicle screw placement.
Cervical Pedicle Screw (CPS) placement is a challenging work due to the high risk of neurovascular complications. Although there have been several different free-hand or navigation assisted techniques for CPS placement, perforations may occur during screw insertion, especially lateral perforation. The objective of this manuscript is to describe a novel free-hand technique for subaxial CPS placement (C3-C7) and to evaluate if it decreases the chances of perforation. Thirty-two patients undergoing surgery with CPS instrumentation (C3-C7) at our institute between June 2017 and December 2018 were included in this study. All the patients had cervical trauma, and pedicle screw insertion was performed according to the free-hand "slide technique". The lamina, lateral mass and facet joint of the target area were exposed and the optimal entry point was found on the lateral mass posterior surface. A pedicular probe was then inserted and gently advanced. During the pedicle probe insertion, the cortex of the medial margin of the pedicle acted as a slide to permit the safe insertion of the screw. If the pedicle screw pathway was intact, the screw of the appropriate size was carefully placed. Three-dimensional (3D) CT imaging reconstruction was performed in all the patients after surgery, and screw perforations were graded with the Gertzbein-Robbins classification. Thirty-two patients who met the inclusion criteria were included in this study. A total of 257 CPSs (C3-7) were inserted, of which 41 CPSs were in C3, 61 CPSs were in C4, 55 CPSs were in C5, 53 CPSs were in C6, and 47 CPSs were in C7. The diameter and length of CPSs were 3.5 mm and 22-26 mm respectively. According to the Gertzbein-Robbins classification, grade 0, 231 screws; grade 1, 19 screws; and grade 2, 7 screws. No neurovascular complications occurred stemming from malpositioning of pedicle screws. Among perforated screws (26 screws), there were 16 lateral perforations, 5 medical perforations, and 4 inferior perforations. The initial usage result shows the "slide technique" is a safe, effective and cost-effective technique for pedicle screw placement in the cervical spine. This is the first report of such a technique, and further studies are needed.


An atypical presentation of lemierre syndrome of urogenital source.
Lemierre syndrome has classically been described as septic thrombophlebitis of the internal jugular veins resulting from primary oropharyngeal disease caused by Fusobacterium necrophorum. However, many cases have been reported of the syndrome originating from other primary sites of infection, resulting in limited utility of the diagnostic criteria and increased difficulty in making the diagnosis. We describe a 22-year-old female who developed the syndrome through a Fusobacterium infection which originated from a Bartholin gland abscess, a phenomenon that has not yet been described in the literature. Understanding and recognizing the variable forms of Lemierre syndrome allows us to better define the condition and provide optimal care to patients.


Preparation of starch-palmitic acid complex nanoparticles and their effect on properties of the starch composite film.
The corn starch-palmitic acid complex nanoparticles and amylose-palmitic acid complex nanoparticles were prepared through complex and nanoprecipitation. Their mean size values were 138.2 nm and 654.7 nm, respectively, while the two kinds of complex nanoparticles were mainly showed V-type crystalline structure, the crystallinity of these complex nanoparticles was 20.86 % and 46.81 %. Then the starch composite films were prepared using the corn starch-palmitic acid complex nanoparticles and amylose-palmitic acid complex nanoparticles as reinforcement phases. The starch composite film reinforced with amylose-palmitic acid complex nanoparticles had the higher tensile strength and a better wettability with the water contact angle of 86.51°. Though the crystalline properties of starch composite films had no significant difference, the thermal stability improved when the amylose-palmitic acid complex nanoparticles used as reinforcement phase, the maximum thermal degradation temperature was 313 °C. This study provides a new type of reinforcement phase to improve the properties of starch composite films.


Synthesis of methyl-branched fatty acids from methylmalonyl-CoA by fatty acid synthase from both the liver and the harderian gland of the guinea pig.
Partially purified fatty acid synthase preparations from both the liver and the harderian gland of guinea pig showed the same relative rates of utilization of methylmalonyl-CoA when compared to malonyl-CoA. Radio gas-liquid chromatographic analysis of the products generated from [methyl-14C]methylmalonyl-CoA and from [2-14C]malonyl-CoA in the presence of unlabeled methylmalonyl-CoA showed that the enzyme from both tissues generated identical mixtures of branched fatty acids. Therefore, it is concluded that the production of methyl-branched acids only by the harderian gland is not due to any unique specificity of the fatty acid synthase of this gland, in contrast to the conclusion reached from results obtained from mass spectrometry of the products generated by crude extracts (Y. Seyama, H. Otsuka, A. Kawaguchi, and T. Yamakawa J. Biochem. 90, 789-797, 1981).


Tocopherol-loaded transfersomes: In vitro antioxidant activity and efficacy in skin regeneration.
Transfersomes were prepared by using different polysorbates (i.e., Tween 20, 40, 60 and 80) and loaded with tocopherol acetate, a naturally-occurring phenolic compound with antioxidant activity. The vesicles showed unilamellar morphology, small size (∼85 nm), low polydispersity index (≤0.27), and high entrapment efficiency, which increased as a function of the length of the Tween fatty acid chain (from 72% to 90%). The long-term stability of the formulations was evaluated by means of the Turbiscan™ technology, which indicated their good stability, irrespective of the Tween used. The vesicles efficiently delivered tocopherol to the skin, and showed biocompatibility in vitro in keratinocytes and fibroblasts. Regardless of the Tween used, the transfersomes were able to protect skin cells from the oxidative damage induced by hydrogen peroxide. Additionally, transfersomes promoted cell proliferation and migration, which resulted in an acceleration of skin wound closure. These results demonstrated that tocopherol-loaded transfersomes bear potential as topical delivery system with antioxidant activity and wound healing properties.


Impact of a mother's weight history on her offspring.
To explore the relationship between maternal lifelong body weight history and anthropometric measurements in the offspring. We studied a prospective sample of 48 pregnant women with either gestational diabetes mellitus (GDM, n = 21) or normal glucose tolerance (NGT, n = 27). Reported maternal weight at birth, 20 years of age and 30 years of age, and pre-pregnancy and maximal weight outside pregnancy were obtained by questionnaire. BMI was calculated using data from the questionnaire. Maternal anthropometric parameters were measured during pregnancy. Offspring anthropometrics were obtained at birth and eight weeks later. Maternal weight at birth, weight or BMI at 20 years of age and at 30 years of age, and maximal weight or BMI did not differ between groups. In all women, maternal birth weight, BMI at 20 years of age, and maximal BMI correlated with newborn birth weight (ρ = 0.39, 0.37, and 0.27, respectively, P ≤ 0.05), with newborn length (ρ = 0.46, 0.32, and 0.30 respectively, P < 0.05), and with infant weight eight weeks later (ρ = 0.43, 0.30, and 0.31, respectively, P < 0.05). Reported maternal BMI at 30 years of age correlated with infant weight (ρ = 0.31) and cranial circumference (ρ = 0.33) at eight weeks of life (P < 0.05). Besides gestational age, maternal weight at screening was the most significant predictor of infant birth weight. Several parameters of maternal weight history were related to offspring anthropometric measurements in early life in a sample of women with and without GDM.


Biological and environmental monitoring of occupational exposure to cyclophosphamide in industry and hospitals.
The aims of the study were to clarify potential exposure situations to anticancer agents during industrial processing, drug manufacture and hospital administration, using cyclophosphamide (CP) as the model compound. CP is considered an animal and human carcinogen, and it is shown to be an indirect mutagen in various test systems using several genetic endpoints. Environmental monitoring was performed by collecting ambient air samples during the different processing and handling stages. Both stationary and personal sampling was used. CP was analyzed by liquid chromatography (HPLC) and mass spectrometry (MS). The process materials and intermediates were also analyzed for genotoxic activity using the Ames test and SCE induction in CHO cells as endpoints. Biological monitoring studies were performed on 147 persons representing 5 groups of workers, control subjects and patients. In the experimental part of the project, the intermediates in the CP manufacturing process, CP I (nor-nitrogen mustard) and CP II (phosphoroxydichloride mustard) were found directly active in the 2 genotoxicity tests. These findings led to improvements in work hygiene when handling CP I and CP II in the process. The CP measurements showed that the highest potential-exposure sites occurred during specific operations of the process, e.g., during emptying of the drying drum and during tablet mass preparation (the range of CP concentrations in air was 0.16-0.49 mg/m3). The correlation between indirect genotoxicity and chemical analyses of the ambient air samples was good, revealing the activity to be due to cyclophosphamide. However, the air samples were found mutagenic without metabolic activation also in the beginning of the process; this is obviously due to CP II particles in the ambient air, since no CP was detected chemically. The personal protection of workers in the plant collaborating in the study is efficient and the production unit is equipped with the best available techniques to protect both the personnel and the quality of the drug. Both the urine mutagenicity analyses using strain TA1535 of Salmonella typhimurium as indicator and the cytogenetic analyses of peripheral blood lymphocytes using sister-chromatid exchanges or structural chromosomal aberrations as endpoints were negative. However, a statistically nonsignificant trend in increased number of micronuclei was observed in binucleated lymphocytes of the worker groups as compared with controls. The studies on the hospital use of CP were performed in 3 oncological units and 1 pharmacy unit.(ABSTRACT TRUNCATED AT 400 WORDS)


Diabetes insipidus associated with propofol anesthesia.
Diabetes insipidus is a clinical entity that is often seen in neurosurgical patients either during or immediately after transsphenoidal hypophysectomy. Rarely, diabetes insipidus can manifest as a new entity months later in patients who have previously had an intracranial injury or operation.


Changes in glycosaminoglycans and proteoglycans of normal breast and fibroadenoma during the menstrual cycle.
Fibroadenoma is the most common breast tumor in young women, and its growth and metabolism may be under hormonal control. In the present paper we described the proteoglycan (PG) composition and synthesis rate of normal breast and fibroadenoma during the menstrual cycle. Samples of fibroadenoma and adjacent normal breast tissue were obtained at surgery. PGs were characterized by agarose gel electrophoresis and enzymatic degradation with glycosaminoglycan (GAG) lyases, and immunolocalized by confocal microscopy. To assess the synthesis rate, PGs were metabolic labeled by 35S-sulfate. The concentration of PGs in normal breast was higher during the secretory phase. Fibroadenoma contained and synthesized more PGs than their paired controls, but the PG concentrations varied less with the menstrual cycle and, in contrast to normal tissue, peaked in the proliferative phase. The main mammary GAGs are heparan sulfate (HS, 71%-74%) and dermatan sulfate (DS, 26%-29%). The concentrations of both increased in fibroadenoma, but DS increased more, becoming 35%-37% of total. The DS chains contained more β-d-glucuronic acid (IdoUA/GlcUA ratios were >10 in normal breast and 2-7 in fibroadenoma). The 35S-sulfate incorporation rate revealed that the in vitro synthesis rate of DS was higher than HS. Decorin was present in both tissues, while versican was found only in fibroadenoma. In normal breast, the PG concentration varied with the menstrual cycle. It was increased in fibroadenoma, especially DS. PGs are increased in fibroadenoma, but their concentrations may be less sensitive to hormonal control.


Facile palladium(0)-catalyzed ring expansion reactions of hydroxy methoxyallenyl cyclic compounds via hydropalladation.
Palladium(0)-catalyzed one-atom ring expansion of various hydroxy methoxyallenyl compounds has been achieved in excellent yields without the use of aryl halides. Hydroxy methoxyallenylisoindolinones, -indanones, and -phthalans have been readily converted to the corresponding isoquinolones, naphthoquinones, and isochromanones in the presence of P(o-tolyl)(3).


Fabrication of cotton textile waste-based magnetic activated carbon using FeCl3 activation by the Box-Behnken design: optimization and characteristics.
Cotton textile waste-based magnetic activated carbon was prepared via simultaneous activation-pyrolysis using FeCl3 as a novel activating agent. The response surface methodology based on the Box-Behnken design method was applied to optimize the preparation parameters and predict the specific surface area of the samples. The optimal activated carbon was obtained at a mass ratio of FeCl3/CTW, activation time and activation temperature of 1.62 : 1, 1 h and 700 °C, respectively. The experimental maximum yield and iodine adsorptive value (32.66% and 714.55 mg g-1) of the resultant carbon were close to that of the predicated response values (34.85% and 783.75 mg g-1), respectively. SEM, N2 adsorption-desorption isotherms, XRD, PPMS, FTIR and pHpzc measurements were conducted to analyze the physicochemical characteristics of the optimal sample. The results showed that the carbon matrix had a high specific surface area of 837.39 m2 g-1 with abundant micropores and acidic surface functional groups, and the saturation magnetization (Ms) was 5.2 emu g-1 due to the formation of Fe3O4. The maximum adsorption of Cr(vi) by the carbon reached 212.77 mg g-1. Furthermore, the addition of FeCl3 lowered the pyrolytic carbonization temperature and inhibited the generation of volatiles in the activation-pyrolysis process. Meanwhile, the formation of Fe2O3 and Fe3O4 derived from FeCl3 was beneficial for the development of vast micropores.


Antisense oligonucleotide-induced block of individual GABAA receptor alpha subunits in cultured visual cortex slices reduces amplitude of evoked inhibitory postsynaptic currents.
Whole cell patch clamp recordings were made in layer II-IV from organotypic slices of rat primary visual cortex, explanted at postnatal day 6 and maintained in a serum-free medium. Neurons evinced current clamp characteristics typical for stellate cells. Between 7 and 21 days in culture, both glutamate- and GABA-mediated postsynaptic currents were observed. Long-term culturing in the presence of a degenerate 15-mer antisense oligonucleotide directed against the transcripts of all alpha subunits genes of the GABAA receptor resulted in a dose dependent reduction of evoked GABA synaptic currents. This reduction was maximal (80%) at 20 microM. A randomized control oligo had no effect. Evoked glutamatergic excitatory postsynaptic currents were unaffected following oligo treatment. A 15-mer antisense oligo directed against the alpha 1 subunit gave variable effects: in some cells the amplitude of evoked GABAergic inhibitory postsynaptic currents (IPSCs) was reduced by 50-75%, while in other cells recorded from the same slices, there was little or no effect. An antisense oligo, directed against the alpha 2 subunit, however, gave a consistent and robust 80% reduction of the amplitude of evoked IPSCs. A 15-mer 3-base mismatch oligo against alpha 2 had no effect. We conclude that the alpha 2 subunit functions in postsynaptic GABAA receptors located on or close to the cell bodies of stellate cells. The role of the alpha 1 subunit is less clear, but this subunit seems spatially differentiated. The in situ antisense oligo technique should provide further insight into the biophysical and pharmacological consequences of the subunit composition of ligand gated channels at functional synapses.


Technique for laparoscopic autonomic nerve preserving total mesorectal excision.
With the introduction of total mesorectal excision (TME) for treatment of rectal cancer, the prognosis of patients with rectal cancer is improved. With this better prognosis, there is a growing awareness about the quality of life of patients after rectal carcinoma. Laparoscopic total mesorectal excision (LTME) for rectal cancer offers several advantages in comparison with open total mesorectal excision (OTME), including greater patient comfort and an earlier return to daily activities while preserving the oncologic radicality of the procedure. Moreover, laparoscopy allows good exposure of the pelvic cavity because of magnification and good illumination. The laparoscope seems to facilitate pelvic dissection including identification and preservation of critical structures such as the autonomic nervous system. The technique for laparoscopic autonomic nerve preserving total mesorectal excision is reported. A three- or four-port technique is used. Vascular ligation, sharp mesorectal dissection and identification and preservation of the autonomic pelvic nerves are described.


Rats fed prolonged high protein diets show an increase in nitrogen metabolism and liver megamitochondria.
Rats were fed diets containing 20, 50 and 80% protein for 14 months. The urea excreted by the rats fed diets containing 50 and 80% protein when compared to rats fed diets containing 20% protein increased ca. 2- and 3-fold, respectively, in ca. 2 days; this increase was maintained essentially unchanged through the experimental period. The serum levels of urea increased 2.5- and 4-fold, respectively, in the first days and were also maintained during the experiment. Glutamate dehydrogenase activity of liver remained unchanged. The five urea cycle enzymes increased with respect to the control values. Orotic acid excretion increased as well as orotidylate decarboxylase and orotate phosphoribosyltransferase, but aspartate transcarbamylase did not. The key amino acids involved in the urea and pyrimidine pathways in liver were also measured; aspartic and glutamic acids and citrulline were increased, and ornithine and arginine did not change with the higher protein intake. In general, no differences were observed between animals fed 50 and 80% protein in their diets. Protein synthesis did not increase with the increase of protein content of the diet. Stereological analysis of ultrathin sections showed that the high protein diet induced a significant increment in the volumetric density, numerical density and size of hepatocyte mitochondria. Moreover, the presence of giant mitochondria, a hundred times larger than normal, was also observed in some periportal hepatocytes of rats fed the 80% protein diet.


Persistent dermal lesions in a patient with previous history of visceral leishmaniasis.
Post-kala-azar dermal leishmaniasis (PKDL) is a complication of visceral leishmaniasis (VL) that most frequently occurs after an episode of VL caused by Leishmania donovani. In this case report, we present a 21-year-old male patient with persistent skin lesions and recurrent visceral leishmaniasis (VL) due to Leishmania infantum. The patient did not respond to multiple lines of anti-leishmanial treatment (including Liposomal amphotericin B and miltefosine) and later died from cerebral lesions presumed to be secondary to persistent VL.


Ionic excitation of a clone of mouse neuroblastoma.
The electrical properties and the possible regulation of these properties were studied by means of intracellular microelectrode recordings in cells of mouse neuroblastoma clone N1E-115. This clone has high levels of tyrosine hydroxylase and regulates this enzyme. Cells treated for 24 h with 4 muM aminopterin followed by at least 5 days in culture developed rhythmic discharge of action potentials when superfused with phosphate-buffered saline containing less than 0.2 mM calcium or less than 0.2 mM calcium and zero potassium. This ionic excitation occurred in no cells at less than 5 days after treatment with aminopterin but at 5 days or more after treatment, 20% of cells responded to low calcium while 52% responded to low calcium and zero potassium. Concomitant with the development of a susceptibility to ionic excitation was an increase in the average resting membrane potential and morphologic maturation. This ionic excitation of cultured mouse neuroblastoma cells may be useful for studying biochemical events associated with repetitive discharge of action potentials.


Nasal and sleep cycle--possible synchronization during night sleep.
Regular cyclic changes in nostril airflow due to a nasal congestion and decongestion are known in literature as nasal cycle. Registration of breathing from each nostril separately gives possibility to registrate moments of alternative change of airflow of nostrils and periods of nasal cycle. This registration during night sleep shows that the length of these periods are about 1.5h, 3.0 h and 4.5h. The length of these periods are multiple of mean length of sleep cycle--about 1.5h. The alternative change of airflow through nostrils occurs through some of REM stages of the sleep. This shows, that during the night sleep becomes synchronization of nasal and sleep cycles in some of the REM phases of sleep. As a result--length of periods of the nasal cycle are one or more length of sleep cycle.


Long-Lasting Impact of Sugar Intake on Neurotrophins and Neurotransmitters from Adolescence to Young Adulthood in Rat Frontal Cortex.
The detrimental impact of fructose, a widely used sweetener in industrial foods, was previously evidenced on various brain regions. Although adolescents are among the highest consumers of sweet foods, whether brain alterations induced by the sugar intake during this age persist until young adulthood or are rescued returning to a healthy diet remains largely unexplored. To shed light on this issue, just weaned rats were fed with a fructose-rich or control diet for 3 weeks. At the end of the treatment, fructose-fed rats underwent a control diet for a further 3 weeks until young adulthood phase and compared with animals that received from the beginning the healthy control diet. We focused on the consequences induced by the sugar on the main neurotrophins and neurotransmitters in the frontal cortex, as its maturation continues until late adolescence, thus being the last brain region to achieve a full maturity. We observed that fructose intake induces inflammation and oxidative stress, alteration of mitochondrial function, and changes of brain-derived neurotrophic factor (BDNF) and neurotrophin receptors, synaptic proteins, acetylcholine, dopamine, and glutamate levels, as well as increased formation of the glycation end-products Nε-carboxymethyllysine (CML) and Nε-carboxyethyllysine (CEL). Importantly, many of these alterations (BDNF, CML, CEL, acetylcholinesterase activity, dysregulation of neurotransmitters levels) persisted after switching to the control diet, thus pointing out to the adolescence as a critical phase, in which extreme attention should be devoted to limit an excessive consumption of sweet foods that can affect brain physiology also in the long term.


A silver-catalyzed stereoselective domino cycloisomerization-vinylogous aldol reaction of ortho-alkynylbenzaldehydes with 3-alkylidene oxindoles: an entry to functionalized isochromenes.
A silver tetrafluoroborate catalyzed domino cycloisomerization-vinylogous aldol addition sequence on a multifunctional substrate such as ortho-alkynylbenzaldehydes yielding functionalized 1H-isochromenes in a single step with high yield and excellent diastereoselectivity (>19 : 1) is described. The reaction was well tolerated by alkyl, aryl, and unsubstituted alkynylbenzaldehydes, and furnished selective 6-endo-dig adducts exclusively without loss in the regio- as well as diastereoselectivity.


The Role of the Gut Microbiome in Colorectal Cancer: Where Are We? Where Are We Going?
Microbiome (microbiota) is a community of all microorganisms inhabiting a specific site of the body, including pathogens, which distinguishes it from the physiological microflora. Intestinal dysbiosis plays a key role in the development of colorectal cancer. In the process of carcinogenesis, inflammation, immune response, and toxic metabolites play a significant role. Specific species of bacteria might affect the risk of colorectal cancer and growth of tumor already present. Assessment of changes in the intestinal microbiome during the development and progression of colorectal cancer might create a simple diagnostic tool, a useful biomarker, or might influence treatment strategies in colorectal cancer patients. Analysis of the gut microbiome provides the potential to develop noninvasive diagnostic tests that would be useful as new protective markers of colorectal cancer, prognostic markers in already present colorectal cancer, and predictive markers of response to treatment, especially immunotherapy.


Electrophysiological characteristics of lumbosacral evoked potentials in patients with established spinal cord injury.
Surface electrodes positioned over the S1 and T12 vertebrae and referenced to T6 were used to record spinal potentials evoked by unilateral stimulation of the posterior tibial nerve at the knee. Data were collected on 24 patients who received spinal cord injuries 2 months to 31 years previously. The recording sites were below the level of spinal injury. The lumbosacral evoked potentials (LSEPs) were compared with the results of measurements obtained from 19 neurologically healthy subjects. Additional data were collected on each patient to characterize segmental reflex responses and preservation of sensory and motor functions associated with the L5 through S2 segments of the spinal cord. Assuming that the LSEP reflects the activity of spinal cord interneurons, the results demonstrate a degree of spinal cord dysfunction caudal to the area of injury in a substantial number of the patients with spinal cord injury which we studied.


HbAlc in an unselected population of 4438 people with type 2 diabetes in a Danish county.
To describe the use and level of HbA1c in a large unselected Type 2 diabetic population in Denmark. In addition, to describe the characteristics of the patients and the general practitioners in relation to the monitoring of HbA1c. Data were collected from public data files for the period January 1993 to December 1997. The County of Vejle with a background population of 342,597 citizens, 303,250 of whom were listed with participating general practitioners. The Type 2 diabetic population alive and resident in the county on 1 January 1997. In a population of 4438 Type 2 diabetics, 73% had a minimum of one annual HbA1c measurement in 1997. No HbA1c measurement was associated with a long history of diabetes, diet treatment or old age. Poor glycaemic regulation was found in 65% of the Type 2 diabetics in 1997. Poor glycaemic regulation was associated with tablet or insulin treatment, age under 70 years and long history of diabetes. The interpractice variation was huge. The quality of HbA1c monitoring of Type 2 diabetics needs to be improved. Possibilities for improvement seem to be present.


Non-curative gastrectomy for metastatic gastric cancer: rationale and long-term outcome in multicenter settings.
Metastatic gastric cancer remains a significant problem as the majority of Western patients are diagnosed with disseminated disease and no routine therapeutic regimen is accepted in such cases. A cohort of 3141 patients with gastric cancer operated between 1990 and 2005 was evaluated using a multicenter data set held by the Polish Gastric Cancer Study Group to determine potential risks and benefits of non-curative gastrectomy for metastatic disease. Additionally, parameters of Quality of Life (QoL) were evaluated prospectively in 140 patients undergoing gastrectomy using the QLQ-C30 questionnaire. Gastrectomy was carried out in 2258 patients. Distant organ metastases were diagnosed in 951 patients, 415 of which underwent non-curative gastrectomy. The overall mortality rates were significantly higher in patients undergoing non-resectional surgery (10%) than either curative (3%, P < 0.001) or non-curative (4%, P = 0.002) gastrectomy. The overall median survival in patients with metastatic disease was significantly higher for non-curative gastrectomy (10.6 months, 95% confidence interval (CI) 9.3-11.9) than for non-resective operations (4.4 months, 95% CI 4.0 to 4.8, P < 0.001). The hazard ratio of death in patients subject to non-resectional surgery compared to those treated by gastrectomy was 2.923 (95% CI 2.473 to 3.454, P < 0.001). A gradual impairment in QoL parameters was found over 12 months after non-curative resections but changes did not reach statistical significance and individual parameters were similar to gastrectomy without distant metastases. Non-curative gastrectomy for metastatic gastric cancer is associated with significantly better survival compared to non-resective surgery and does not impair quality of life.


Frequency and predictors of implant-related complications after posterior cervical fusion.
Retrospective, observational study. To determine the frequency and predictors of implant-related complications in adults after posterior cervical fusion. Published literature on lumbosacral fusion suggest that implant-related complications are not uncommon. Although posterior cervical fusion is a common operation, data on frequency and predictors of implant-related complications after posterior cervical fusion is still scarce. 86 patients (with 740 screws) who underwent posterior cervical fusion were included. Implant-related complications were identified by the presence of: (1) halo sign, (2) screw pull-out/breakage (3) post-operative kyphosis and (4) implant-related complications requiring revision surgery. These were stratified into two groups: (a) minor - isolated halo sign or screw pull-out/breakage (b) major - post-operative kyphosis > 10 degrees, and revision surgery. Demographic, operative and radiological data was collected. Rates of implant-related complications were determined and associated risk factors identified. 33 (38.4%) patients had signs of implant-related complications. Of these, 29 (87.9%) had minor complications and 4 (12.1%) had major complications. Charlson Comorbidity Index (CCI) (p = 0.03179) and pre-op C2-C7 sagittal vertical alignment (SVA) (p = 0.02449) were the only significant risk factors for all-cause implant-related complications during multivariate logistic regression. Other intraoperative parameters (type of screw, length of fusion, levels decompressed, and extension of fusion beyond the levels decompressed) were not significantly associated with implant-related complications. Implant-related complications are not uncommon but rarely require revision surgery. Higher pre-operative SVA and CCI were significant risk factors; length of construct and extent of decompression were not. These findings may assist clinicians when deciding the extent of fusion and in selecting patients for closer follow-up.


Fully automated interferon-γ release assay to monitor antigen-independent T cell functionality: A proof of concept study in septic shock.
In sepsis, personalized immunotherapy is being evaluated as a means of restoring immune function in the most severely affected patients. Biomarkers play a crucial role in this process, as there are no clear clinical indicators of immune dysfunction. Functional testing is considered a gold standard for assessing immune function, but this approach faces analytical challenges in terms of clinical implementation. The use of technician-dependent, time-consuming, home-made protocols often leads to poor standardization. This study represents the first beta testing of a fully automated interferon-γ release assay (IGRA) for monitoring the functionality of antigen-independent T lymphocytes. We observed a significant decrease in IFN-γ release capacity, which was associated with typical alterations in immunological cellular parameters (such as low mHLA-DR expression and decreased CD8 T lymphocyte count), in 22 patients with septic shock. Since the test is performed using whole blood and requires no technician intervention, with results available within 4 h, it may offer new possibilities for monitoring patients with immune alterations in routine clinical conditions. Further investigations in larger cohorts of patients are now needed to validate its clinical potential.


Recent advances in the study of biomolecular interactions by capillary electrophoresis.
Capillary electrophoresis (CE) has been abundantly used in the study of molecular interactions owing to such advantages as short analysis time, low sample size requirement, high separation efficiency, and flexible applications. The focus of this paper is to review recent studies and advances (mainly from 1998 to now) in biomolecular interactions using CE. Five CE modes: zone migration CE, affinity CE, frontal analysis (FA), Hummel-Dreyer (HD) and vacancy peak (VP) are cited and compared. Quantitative aspects of the thermodynamics and kinetics of biomolecular interaction are reviewed. Several biomolecular binding systems, including protein-protein (polypeptide), protein-DNA (RNA), protein(polypeptide)-carbohydrate, protein-small molecule, DNA-small molecule, small molecule-small molecule, have been well characterized by CE. CE is shown to be a powerful tool for the determination of the binding parameters of various bioaffinity interactions.


HB-EGF is a paracrine growth stimulator for early tumor prestages in inflammation-associated hepatocarcinogenesis.
We studied the impact of heparin-binding epidermal growth factor-like growth factor (HB-EGF) on inflammation-driven hepatocarcinogenesis. HB-EGF expression was determined by qRT-PCR and immunodetection in hepatocellular adenoma and carcinoma and in mesenchymal (MC) and parenchymal liver cells obtained from different models of inflammation. The functions of HB-EGF in early hepatocarcinogenesis were assessed in co-cultures of unaltered and initiated/premalignant hepatocytes. In human and rat (pre)malignant liver lesions, HB-EGF levels were comparable to that of the surrounding tissue. In inflamed livers HB-EGF was expressed predominantly in MC and was further increased by pro-inflammatory lipopolysaccharide (LPS) or linoleic acid hydroperoxide (LOOH). In culture, DNA-replication occurred rather in initiated/premalignant than unaltered hepatocytes and was further elevated by LOOH- or LPS-stimulated MC-supernatants. The supernatant effects were abrogated by pre-incubation with HB-EGF-neutralizing antisera. HB-EGF itself induced DNA-replication and mitosis preferentially in the initiated/premalignant cells. When transducing hepatocytes with a dominant-negative ErbB1-construct, HB-EGF-induced DNA-replications were blocked completely in unaltered hepatocytes but incompletely in initiated/premalignant cells, which suggests elevated ErbB-mediated signal transduction in first stages of hepatocarcinogenesis. Pro-inflammatory stimuli induce the release of HB-EGF from MC, which stimulates DNA-replication in initiated/premalignant hepatocytes. Similar mechanisms may contribute to carcinogenesis in human inflammatory liver diseases.


Use of the extensor retinaculum as a donor site for bone-ligament-bone grafts.
The recent introduction of bone-ligament-bone grafts for ligament reconstruction in hand surgery has led to a search for suitable graft donor sites. The extensor retinaculum had been suggested, but harvesting details were still lacking. We therefore studied the anatomy of the extensor retinaculum, with radiography and morphometric measurements. Ten cadaver wrists were dissected; the thick portion of the retinaculum and its insertions on the radius in the different extensor compartments were studied. The compartments were wire-marked, and AP radiographs taken of each specimen. The retinaculum spanning the first three compartments was removed with the bony ridges it inserted on, and the inter-ridge retinaculum length was measured. A comparison of our dissections with the radiographs showed that only the radial compartments 1, 2 and 3 had constant bony insertions on both ridges, making them suitable for bone-ligament-bone grafting. The mean compartment length was 11.7 mm (compartment 1), 17 mm (compartment 2) and 7.3 mm (compartment 3) respectively. Thus, the extensor retinaculum can be relied upon to provide bone-ligament-bone grafts from the first three compartments, for possible use in ligament reconstructions in the hand and wrist.


Effects of selenium levels on placental oxidative stress and inflammation during pregnancy: a prospective cohort study.
Studies on the impact of Se levels in different pregnancy periods on placental function are limited. This cohort study sought to investigate the levels of the trace element Se and to assess their effects on placental oxidative stress (OS) and mRNA expression of inflammatory genes during pregnancy. The study population consisted of 2519 pregnant women from the Ma'anshan birth cohort. Se levels were measured in the first and second trimesters of pregnancy and in cord blood using inductively coupled plasma-mass spectrometry (ICP-MS). Placental stress and mRNA expression of inflammatory genes were assessed using RT-PCR. A statistically significant negative association was noted between Se levels in the second trimester of pregnancy and mRNA expression of placental HO-1(β = -0.009, p < .01), HIF1α (β = -0.005, p = .010), GRP78 (β = -0.011, p < .001), CRP (β = -.007, p = .033) and CD68 (β = -0.006, p = .019). A negative association was noted between Se levels in cord blood and mRNA expression of placental HO-1 (β = -0.007, p = .004), HIF1α (β = -0.006, p = .005) and GRP78 (β = -0.009, p = .004). We found that prenatal Se status was associated with placental stress and mRNA expression of inflammatory genes. Se deficiency during pregnancy, especially in the second trimester, leads to the production of OS and an increase in inflammatory mediators, affecting the growth and development of the fetus. Monitoring of pregnant women's nutritional status is necessary to prevent nutritional imbalances and deficiencies in important micronutrients in the fetal.


A reweighting of receptor mechanisms in the ground squirrel retina: PIII and B-wave spectral sensitivity functions.
PIII and b-wave spectral sensitivity functions were measured under various adapted conditions in the 13-lined ground squirrel. The PIII was isolated by intravitreal injection of sodium L-aspartate. Under the conditions studied, the PIII spectral sensitivity function always fitted the absorption spectrum of a 518 +/- 3 mm Dartnall nomogram. This suggests that of the three photoreceptors ('rod-like', 'blue' cone and 'green' cone) present in the 13-lined ground squirrel the green cones are the overwhelming majority. The dark-adapted b-wave had two components; a b1-wave with a spectral sensitivity that fitted a 516 +/- 1 nm Dartnall nomogram and a b2-wave with a spectral sensitivity that fitted a 502 +/- 4 nm Dartnall nomogram. The light-adapted b-wave had a spectral sensitivity that fitted a 516 +/- 2 nm Dartnall nomogram.


Intracranial blood velocity in head injury. A transcranial ultrasound Doppler study.
We report our experience monitoring head-injured patients by means of transcranial Doppler sonography. Cerebral velocity measurements and waveforms change in a consistent pattern with the existence of intracranial pressure, and it is possible to discriminate low versus high flow states. The technique, in contrast to cerebral blood flow measurement, is reliable, atraumatic, and repeatable so that diagnostic assessments can be made and the patient can be followed for therapeutic efficacy.


How much can we trust electronic health record data?
Trust in EHR data is becoming increasingly important as a greater share of clinical and health services research use EHR data. We discuss reasons for distrust and acknowledge limitations. Researchers continue to use EHR data because of strengths including greater clinical detail than sources like administrative billing claims. Further, many limitations are addressable with existing methods including data quality checks and common data frameworks. We discuss how to build greater trust in the use of EHR data for research, including additional transparency and research priority areas that will both enhance existing strengths of the EHR and mitigate its limitations.


PD-1 has a unique capacity to inhibit allergen-specific human CD4+ T cell responses.
T lymphocytes have a crucial role in initiating and promoting type I allergies. Their responses are tightly regulated by numerous activating and inhibitory signals provided by APCs. Here we have addressed the role of the major coinhibitory receptors PD-1, CTLA-4, BTLA and LAG-3 in allergen-specific CD4+ T cell responses. PBMCs of healthy individuals and 41 patients allergic to house dust mites, birch, grass or mugwort pollen were stimulated with allergenic extracts and expression of coinhibitory receptors on responding CD4+ T cells was assessed. Blocking antibodies to PD-1, CTLA-4, BTLA and LAG-3 were used to evaluate the role of coinhibitory pathways. Allergen-specific CD4+ T cells showed strong upregulation of PD-1, LAG-3 and CTLA-4 upon stimulation, whereas BTLA was downregulated. Blockade of PD-1 strongly enhanced proliferation and cytokine production (IL-10; TH1 cytokines IFN-γ, TNF-α; TH2 cytokines IL-5, IL-13) of allergen-specific CD4+ T cells derived from allergic as well as non-allergic individuals. BTLA blockade enhanced proliferation but not cytokine production in response to house dust mite extract. Blocking LAG-3 was ineffective and surprisingly, we observed reduced proliferation and cytokine production in presence of a CTLA-4 antibody. Our results point to a unique potency of PD-1 pathways to dampen allergen-specific human T cells.


Evaluation of the intraoperative specimens of the thoracic and abdominal aorta.
Little is known about the histological patterns of acute and chronic aortic pathology with regard to medial degeneration, atherosclerosis and aortitis as well as their distribution in different age groups. The aim of the study was to evaluate histopathological findings of intraoperatively gained aortic specimens with regard to the incidence of medial degeneration, atherosclerosis and aortitis. Intraoperatively gained aortic specimens were evaluated in 151 patients including 83 (55%) aortic aneurysms (65 thoracic, 18 abdominal) and 68 (45%) acute type A aortic dissections. Histological stainings used were hematoxylin and eosin, Van Gieson as well as alcian blue. Patients were stratified according to above and below 65 years of age. High grade medial degeneration represented pooling of mucoid material in the whole aortic wall. High grade atherosclerosis represented severe intimal fibrosis, massive accumulation of macrophages and foam cells or massive calcification of the aortic wall. Medial degeneration was diagnosed in 106 (70%) patients including 55 (52%) aortic aneurysms and 51 (48%) acute type A aortic dissections. High grade medial degeneration was found in 50% of patients with thoracic aortic aneurysms < 65 years of age vs. 44% in patients ≥ 65 years of age (p = 0.64) and in 36% of patients with thoracic aortic dissections < 65 years of age vs. 14% in patients ≥ 65 years of age (p = 0.07). Atherosclerosis was diagnosed in 71 (47%) patients including 46 (65%) aortic aneurysms and 25 (35%) aortic dissections. High grade atherosclerosis was found in 23% of patients with thoracic aneurysms < 65 years of age vs. 36% in patients ≥ 65 years of age (p = 0.24) and in 13% of patients with aortic dissections < 65 years of age vs. 52% in patients ≥ 65 years of age (p < 0.001). Aortitis was rare (n = 2). Medial degeneration was the most frequent diagnosis in this series of aortic specimens. Medial degeneration was equally common in patients above and below 65 years of age. However in cases with acute type A aortic dissections, high grade atherosclerosis was the leading histopathological diagnosis in patients older than 65 years. Acute type A aortic dissections seem to have different underlying pathologies in different age groups.


Anterior pituitary function before and after trans-sphenoidal microsurgical resection of pituitary tumors.
Anterior pituitary lobe function was measured pre- and postoperatively in 40 patients with pituitary tumors who were managed surgically by the trans-sphenoidal approach. Of 23 patients with normal anterior lobe function preoperatively, 21 (91%) were normal postoperatively. Of 17 preoperatively impaired patients, 7 (40%) regained normal function, 3 (18%) improved, 4 (24%) remained the same, and 3 (18%) were further impaired by the surgery. In this small series, the likelihood of recovery of pituitary function varied inversely with the degree of preoperative impairment and the size of the tumor, suggesting that more aggressive surgical management of small, minimally symptomatic pituitary tumors is justified.


Predicting program attendance and weight loss in obesity interventions: Do triggering events help?
Medical events that "trigger" motivation to lose weight may improve treatment outcomes compared to non-medical or no triggering events. However, previous findings include only long-term successful participants, not those initiating treatment. The current study compared those with medical triggering events or non-medical triggering events to no triggering events on attendance and weight loss during a weight management program. Medical-triggering-event participants lost 1.8 percent less weight (p = 0.03) than no-triggering-event participants. Non-medical-triggering-event participants attended 1.45 more sessions (p = 0.04) and were 1.83 times more likely to complete the program (p = 0.03) than no-triggering-event participants. These findings fail to support the benefit of medical triggering events when beginning treatment for obesity.


[Development of radioimmunoassay for human sperm protein 17].
To develop a method of radioimmunoassay for human sperm protein 17(Sp17) determination. Anti-recombinant human Sp17 antibody was prepared, the labeling of 125I-rhSp17 performed by chloramine T method, and radioimmunoassay of Sp17 developed. The assay range was 3.3 to approximately 800 microg/L, the sensitivity was 2.0 microg/L, and the intra- and inter-assay coefficients of variation (CV) were 7.5% to approximately 9.8% and 8.2% to approximately 13.2%, respectively. The serum Sp17 level in normal subjects was (15.60 +/- 7.66) microg/L (n = 59). This radioimmunoassay of Sp17 fulfills the reasonable requirements of clinical routine and scientific studies in terms of specificity, sensitivity and practicability. Measurement of Sp17 concentration is useful for assessing its native distribution and aberrant expression.


Mass spectrometry-based metabolite profiling and antioxidant activity of Aloe vera ( Aloe barbadensis Miller) in different growth stages.
Metabolite profiling of four different-sized Aloe vera plants was performed using gas chromatography-ion trap-mass spectrometry (GC-IT-MS) and ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS) with multivariate analysis. Amino acids, sugars, and organic acids related to growth and development were identified by sizes. In particular, the relative contents of glucose, fructose, alanine, valine, and aspartic acid increased gradually as the size of the aloe increased. Anthraquinone derivatives such as 7-hydroxy-8-O-methylaloin, 7-hydroxyaloin A, and 6'-malonylnataloins A and B increased gradually, whereas chromone derivatives decreased continuously as the size of the aloe increased. The A30 aloe (size = 20-30 cm) with relatively high contents of aloins A and B, was suggested to have antioxidant components showing the highest antioxidant activity among the four different sizes of aloe. These data suggested that MS-based metabolomic approaches can illuminate metabolite changes associated with growth and development and can explain their change of antioxidant activity.


Possible role of adipocytokines in systemic sclerosis-associated small pericardial effusion.
Pericardial effusion is a common manifestation of systemic sclerosis, but its pathogenesis has been poorly investigated. Adipokines and interleukins may play a role in the pathophysiology of pericardial effusion. This study aimed at evaluating serum levels of adipokines and interleukins in systemic sclerosis patients with and without pericardial effusion. A total of 87 systemic sclerosis patients (age 52.6 ± 14 years; disease duration 8.2 ± 6.7 years) were recruited in this study. Demographics, body mass index, and clinical characteristics were recorded in each patient. Pericardial effusion was considered pathologic when ≥50 mL was detected by echocardiography. Serum levels of adiponectin, leptin, resistin, visfatin, tumor necrosis factor-α, interferon-γ, interlueukin-2, interlueukin-10, and interlueukin-17 were measured using Multiplex Immunoassay (Bioplex 200 System). In all, 11 (13%) systemic sclerosis patients had pericardial effusion. Systemic sclerosis patients with and without pericardial effusion did not differ in age, sex, and body mass index. Systemic sclerosis patients with pericardial effusion had significantly higher levels of visfatin (median/interquartile range: 1546 pg/mL (interquartile range: 8590) vs 388 pg/mL (interquartile range: 103), p = 0.03) and interlueukin-17 (1.33 pg/mL (interquartile range: 3.5) vs 0.05 pg/mL (interquartile range: 0.56), p = 0.04), but lower levels of adiponectin (2,845,000 pg/mL (interquartile range: 4,132,900) vs 5,272,100 pg/mL (interquartile range 8,243,600), p = 0.02) than patients without pericardial effusion. Interstitial lung disease, pulmonary arterial hypertension, and "limited" or "diffuse" cutaneous subset did not correlate to adipokines or interleukin levels. Visfatin and adiponectin may play an important role in the pathogenesis of systemic sclerosis-related pericardial effusion. Further longitudinal studies are needed to unravel a possible role of these molecules as biomarkers of pericardial effusion in systemic sclerosis patients.


Japanese guidelines for childhood asthma 2020.
The Japanese Guideline for Childhood Asthma (JGCA) 2020 is a translation of the Japanese Pediatric Guideline for the Treatment and Management of Asthma (JPGL) 2017 into English, which was published by the Japanese Society of Pediatric Allergy and Clinical Immunology. It makes recommendations for best practices in the management of childhood asthma, including management of acute exacerbations and non-pharmacological and pharmacological management. These guidelines will be of interest to non-specialist physicians involved in the care of children with asthma. In JPGL, JPGL2017 is the first evidence-based guidelines updated according to the GRADE system and Minds approach, and it addresses eight clinical questions about the treatment of childhood asthma. In children aged ≤5 years, infant and preschool asthma is diagnosed according to the response to short acting beta2 agonists or the effect of a therapeutic trial during 1 month with controller treatment and worsening after treatment cessation. Long-term management both promotes pharmacological therapy and measures against risk factors that induce exacerbation, better patient education and a partnership with trinity. In addition, long-term management should not be carried out without review but rather be based on a cycle of evaluation, adjustment and treatment. In JPGL2017, the transdermal patch and oral beta2 agonists are positioned as drugs within the concept of "short-term additional treatment" to be used until the symptoms are stabilized when the control state transiently deteriorates.


Vascular anomalies - a practical approach.
Vascular anomalies are common clinical problems (around 4.5% of all patients) in pediatric dermatology. A correct diagnosis is possible on clinical grounds in around 90% of cases; the remaining patients may require radiologic evaluations (duplex ultrasonography, MRI scan) and, rarely, histology. Vascular anomalies are divided into tumors and vascular malformations. This clear division reflects the different biological behaviors of these two groups. The infantile hemangioma represents by far the most common vascular tumor and is characterized by a typical growth cycle consisting of rapid proliferation, plateau phase, and finally slow regression. The discovery in 2008 of the efficacy of beta blockers in this disease is a therapeutic milestone. Vascular malformations can affect all types of vessels (capillaries, veins, arteries and lymphatic vessels). They usually manifest at birth and grow proportionally with the affected child. Some show marked progression especially during puberty. Considerable progress has been made with innovative interventional therapies in recent years, but surgery remains an important option. Basic knowledge of these diseases is important to every dermatologist in order to be able to counsel and manage affected patients correctly.


Graphene Supported Rhodium Nanoparticles for Enhanced Electrocatalytic Hydrogen Evolution Reaction.
Current research on catalysts for proton exchange membrane fuel cells (PEMFC) is based on obtaining higher catalytic activity than platinum particle catalysts on porous carbon. In search of a more sustainable catalyst other than platinum for the catalytic conversion of water to hydrogen gas, a series of nanoparticles of transition metals viz., Rh, Co, Fe, Pt and their composites with functionalized graphene such as RhNPs@f-graphene, CoNPs@f-graphene, PtNPs@f-graphene were synthesized and characterized by SEM and TEM techniques. The SEM analysis indicates that the texture of RhNPs@f-graphene resemble the dispersion of water droplets on lotus leaf. TEM analysis indicates that RhNPs of <10 nm diameter are dispersed on the surface of f-graphene. The air-stable NPs and nanocomposites were used as electrocatalyts for conversion of acidic water to hydrogen gas. The composite RhNPs@f-graphene catalyses hydrogen gas evolution from water containing p-toluene sulphonic acid (p-TsOH) at an onset reduction potential, Ep, -0.117 V which is less than that of PtNPs@f-graphene (Ep, -0.380 V) under identical experimental conditions whereas the onset potential of CoNPs@f-graphene was at Ep, -0.97 V and the FeNPs@f-graphene displayed onset potential at Ep, -1.58 V. The pure rhodium nanoparticles, RhNPs also electrocatalyse at Ep, -0.186 V compared with that of PtNPs at Ep, -0.36 V and that of CoNPs at Ep, -0.98 V. The electrocatalytic experiments also indicate that the RhNPs and RhNPs@f-graphene are stable, durable and they can be recycled in several catalytic experiments after washing with water and drying. The results indicate that RhNPs and RhNPs@f-graphene are better nanoelectrocatalysts than PtNPs and the reduction potentials were much higher in other transition metal nanoparticles. The mechanism could involve a hydridic species, Rh-H- followed by interaction with protons to form hydrogen gas.


Genes and experience shape brain networks of conscious control.
One aspect of consciousness involves voluntary control over thoughts and feelings, often called will. Progress in neuroimaging and in sequencing the human genome makes it possible to think about voluntary control in terms of a specific neural network that includes midline and lateral frontal areas. A number of cognitive tasks involving conflict as well as the control of emotions have been shown to activate these brain areas. Studies have traced the development of this network in the ability to regulate cognition and emotion from about 2.5 to 7 years of age. Individual differences in this network have been related to parental reports of the ability of children to regulate their behavior, to delay reward and to develop a conscience. In adolescents these individual differences predict the propensity for antisocial behavior. Differences in specific genes are related to individual efficiency in performance of the network, and by neuroimaging, to the strength of its activation of this network. Future animal studies may make it possible to learn in detail how genes influence the common pattern of development of self-regulation made possible by this network. Moreover, a number of neurological and psychiatric pathologies involving difficulties in awareness and volition show deficits in parts of this network. We are now studying whether specific training experiences can influence the development of this network in 4-year-old children and if so, for whom it is most effective. Voluntary control is also important for the regulation of conscious input from the sensory environment. It seems likely that the same network involved in self-regulation is also crucial for focal attention to the sensory world.


Time-resolved laser-induced fluorescence spectroscopy for enhanced demarcation of human atherosclerotic plaques.
We report on the enhanced demarcation between human atherosclerotic plaques and normal vessel wall obtained using time-resolved detection of laser-induced fluorescence rather than the customary time-integrated monitoring technique. A frequency-doubled mode-locked and cavity-dumped continuous wave dye laser was used for picosecond pulse generation at 320 nm, and photon-counting techniques were employed for the time-resolved signal monitoring from human aorta samples in vitro. Implications for imaging fluorescence angioscopy and spectroscopic guidance in laser ablation of plaque are indicated.


Human ovarian cancer xenografts in nude mice: chemotherapy trials with paclitaxel, cisplatin, vinorelbine and titanocene dichloride.
The new cytostatics titanocene dichloride and vinorelbine were compared to cisplatin and paclitaxel using a human ovarian cancer xenografts model. Biopsy material from a native human ovarian carcinoma was expanded and transplanted into 96 nude mice. The animals were divided into six treatment groups: cisplatin 3 x 4 mg/kg, paclitaxel 5 x 26 mg/kg, vinorelbine 1 x 20 mg/kg, titanocene dichloride 3 x 30 mg/kg, titanocene dichloride 3 x 40 mg/kg and a control group treated with 0.9% saline. Each experiment was repeated with eight mice in each treatment group. Treatment groups were evaluated in terms of average daily increase in tumor volume and average daily body weight increase of nude mice based on slopes of least-square regressions performed on individual animals. The slope factors alpha and beta of the body weight (alpha) and tumor volume changes (beta) within each group during the course of an experiment were calculated. Both a statistically significant decrease (p<0.05) in the body weight of the experimental animals (cisplatin: alpha = -0.5163, vinorelbine: alpha = -0.6598, paclitaxel: alpha = -0.6746, titanocene dichloride 3 x 30 mg/kg: alpha = -0.6259, titanocene dichloride 3 x 40 mg/kg: alpha = -0.7758) and a significant reduction (p<0.05) of the increase in tumor volume (cisplatin: beta = 12.049, vinorelbine: beta = 0.504, paclitaxel: beta = -1.636, titanocene dichloride 3 x 30 mg/kg: beta = 6.212, titanocene dichloride 3 x 40 mg/kg: beta= -0.685) was shown in all treated groups compared to the control group (alpha = -0.1398; beta = 23.056). No significant weight changes were observed between the individually treated groups. A statistically significant reduction of the tumor growth occured under paclitaxel (beta = -1.636), vinorelbine (beta = 0.504) and titanocene dichloride medication 3 x 40 mg/kg (beta = -0.685), as compared to the group treated with cisplatin (beta = 12.049). We found titanocene dichloride to be as effective as paclitaxel and more effective than cisplatin. Vinorelbine seems to be a very effective antineoplastic agent exhibiting a significant higher cytostatic effect than cisplatin. Both titanocene dichloride and vinorelbine provide new therapeutic options in women with ovarian carcinoma not responding to standard chemotherapy.


Morphine tissue levels and reduction of gastrointestinal transit in rats. Correlation supports primary action site in the gut.
Overnight-fasted male rats given a single dose of tritium-labeled morphine either intraperitoneally or intravenously were fed a charcoal test meal by stomach tube. The drug remaining in tissues was assayed by liquid scintillation counting of thin-layer chromatograms from homogenates, and gastrointestinal transit was tested by measuring the portion of the small intestine traversed by charcoal in 5 min. Morphine, 0.15 mg/kg, given intraperitoneally either 10 min or 30 min before testing substantially reduced gastrointestinal transit (to 23% and 55% of drug-free controls, respectively), and produced maximum drug levels 5 min after administration in small intestine longitudinal muscle with attached myenteric plexus (500 +/- 42 ng/g, mean +/- SE, n = 4). Intact small intestine, plasma, and brain, respectively, contained decreasing drug concentrations that, in the latter, never exceeded 2%-3% of that in longitudinal muscle. Rats receiving 0.15 mg/kg morphine intravenously presented only minor and short-lived inhibition of gastrointestinal transit that was significantly below (approximately 35%) that of drug-free controls at 10 min, but not 30 min, after drug administration. Morphine levels in the brain and plasma of these rats were up to five times higher, and in the intact small intestine longitudinal muscle were up to 20 times lower than in intraperitoneally treated rats. Morphine concentration in the tissues assayed was plotted against the effect on gastrointestinal transit at the same interval for individual rats regardless of dose, administration route, and observation time: data analysis, in small intestine longitudinal muscle, but not in the brain or plasma, indicated a highly significant correlation and fitting of computer-generated curves described by a currently accepted equation according to the receptor occupation theory of drug response. In view of these findings, and of the complete prevention by the "peripherally selective" narcotic antagonist N-methyl naloxone of gastrointestinal transit inhibition after an intravenous analgesic dose of morphine (1 mg/kg), the investigated animal model is consistent with the primary role of a gut-located action site in opiate-induced constipation.


Reducing oil droplet sizes from a subsea oil and gas release by water jetting a laboratory study performed at different scales.
The main objective of subsea mechanical dispersion (SSMD) is to reduce the oil droplet sizes from a subsea oil release, thereby influencing the fate and behaviour of the released oil in the marine environment. Subsea water jetting was identified as a promising method for SSMD and imply that a water jet is used to reduce the particle size of the oil droplets initially formed from the subsea release. This paper presents the main findings from a study including small-scale testing in a pressurised tank, via laboratory basin testing, to large-scale outdoor basin testing. The effectiveness of SSMD increases with the scale of the experiments. From a five-fold reduction in droplet sizes for small-scale experiments to more than ten-fold for large-scale experiments. The technology is ready for full-scale prototyping and field testing. Large-scale experiments performed at Ohmsett indicate that SSMD could be comparable to subsea dispersant injection (SSDI) in reducing oil droplet sizes.


Recent advances in artificial enzyme cascades for the production of value-added chemicals.
Enzyme cascades are efficient tools to perform multi-step synthesis in one-pot in a green and sustainable manner, enabling non-natural synthesis of valuable chemicals from easily available substrates by artificially combining two or more enzymes. Bioproduction of many high-value chemicals such as chiral and highly functionalised molecules have been achieved by developing new enzyme cascades. This review summarizes recent advances on engineering and application of enzyme cascades to produce high-value chemicals (alcohols, aldehydes, ketones, amines, carboxylic acids, etc) from simple starting materials. While 2-step enzyme cascades are developed for versatile enantioselective synthesis, multi-step enzyme cascades are engineered to functionalise basic chemicals, such as styrenes, cyclic alkanes, and aromatic compounds. New cascade reactions have also been developed for producing valuable chemicals from bio-based substrates, such as ʟ-phenylalanine, and renewable feedstocks such as glucose and glycerol. The challenges in current process and future outlooks in the development of enzyme cascades are also addressed.


Corrigendum to: The long non-coding RNA SPRIGHTLY and its binding partner PTBP1 regulate exon 5 skipping of SMYD3 transcripts in group 4 medulloblastomas.
[This corrects the article DOI: 10.1093/noajnl/vdac120.].


Use of RNA interference in Drosophila S2 cells to identify host pathways controlling compartmentalization of an intracellular pathogen.
Three genome-wide RNA interference screens were performed in Drosophila S2 cells to dissect the contribution of host processes to Listeria monocytogenes entry, vacuolar escape, and intracellular growth. Among the 116 genes identified, several host pathways previously unrecognized as playing a role in listerial pathogenesis were identified: knockdowns affecting vacuolar trafficking to and from the multivesicular body bypassed the requirement for the essential pore-forming toxin listeriolysin O in mediating escape from phagocytic vacuoles and knockdowns affecting either subunit of serine palmitoyltransferase, a key enzyme in ceramide and sphingolipid biosynthesis, enhanced the toxicity of listeriolysin O expressed in the host cell cytosol, leading to lack of appropriate toxin activity compartmentalization and host cell death. Genome-wide RNA interference screens using Drosophila S2 cells proved to be a powerful approach to dissect host-pathogen interactions.


A Study to Evaluate Doravirine Pharmacokinetics When Coadministered With Acid-Reducing Agents.
Doravirine is a novel non-nucleoside reverse transcriptase inhibitor indicated for the treatment of human immunodeficiency virus type 1 infection. Because of potential concomitant administration with acid-reducing agents, a drug-interaction trial was conducted to evaluate the potential impact of these types of medications on doravirine pharmacokinetics. In an open-label, 3-period, fixed-sequence trial, healthy adult participants received the following: period 1, a single dose of doravirine 100 mg; period 2, coadministration of a single dose of doravirine 100 mg and an antacid (1600 mg aluminum hydroxide, 1600 mg magnesium hydroxide, and 160 mg simethicone); period 3, 40 mg pantoprazole once daily on days 1-5 coadministered with a single dose of doravirine 100 mg on day 5. There was a minimum 10-day washout between periods. Plasma samples for pharmacokinetic evaluation were collected, and safety was assessed. Fourteen participants (8 male, 6 female) were enrolled, and 13 completed the trial. Geometric mean ratios (90% confidence intervals) for doravirine AUC0-inf , Cmax , and C24 for doravirine + antacid/doravirine were 1.01 (0.92-1.11), 0.86 (0.74-1.01), and 1.03 (0.94-1.12), respectively, and for doravirine + pantoprazole/doravirine were 0.83 (0.76-0.91), 0.88 (0.76-1.01), and 0.84 (0.77-0.92), respectively. Doravirine was generally well tolerated administered alone or with either of the acid-reducing agents. Coadministration of an aluminum/magnesium-containing antacid or pantoprazole did not have a clinically meaningful effect on doravirine pharmacokinetics, supporting the use of acid-reducing agents with doravirine.


5-Azacytidine induction of stable mesodermal stem cell lineages from 10T1/2 cells: evidence for regulatory genes controlling determination.
5-Azacytidine converts the mouse embryonic cell line C3H 10T1/2 into differentiated chondrocytes, adipocytes, and skeletal muscle. Clonal and 2D protein gel analyses demonstrate that 5-azacytidine converts 10T1/2 cells into three stably determined, but undifferentiated, stem cell lineages which can differentiate into myofibers, chondrocytes, and adipocytes. Conversion of 10T1/2 cells is accompanied by specific changes in protein synthetic patterns unique for each cell lineage. We propose that 5-azacytidine converts 10T1/2 cells by hypomethylation of "determination" regulatory loci which establish lineages of stem cells with a restricted potential to differentiate into muscle, cartilage, or fat cells. Our results suggest that these three lineages are specified by separate regulatory loci and that as few as 1-3 hypomethylation events per cell are sufficient to activate the hypothesized muscle regulatory locus. Conversion of 10T1/2 cells by 5-azacytidine provides a model for studying regulatory genes involved in cell lineage determination.


Effects of regular physical activity on defecation pattern in middle-aged patients complaining of chronic constipation.
It is not well known whether physical activity (PA) is useful in the management of patients complaining of constipation. The aim of this study was to test the influence of regular PA on colonic transit time and defecation in middle-aged inactive patients suffering from chronic idiopathic constipation. Forty-three subjects (> 45 years) were randomly divided into group A (n = 18, 16 F, 2 M) and group B (n = 25, 20 F, 5 M). Group A subjects maintained their normal lifestyle during 12 weeks, followed by a 12-week PA programme. Group B performed a 12-week PA programme after randomization. PA comprised 30 min of brisk walking and a daily 11-min home-based programme. Both groups received dietary advice. Colonic transit time was measured using a radiographic multiple marker single film technique. Despite dietary advice, mean fibre and fluid intake did not change. In group B a significant reduction in 3 out of 4 of the Rome I criteria for constipation was observed, i.e. percentage of incomplete defecations, percentage of defecations requiring straining and percentage of hard stools (p < 0.05). As a consequence, the number of fulfilled Rome criteria for constipation decreased (2.7 to 1.7; p < 0.05). Furthermore, the rectosigmoid and total colonic transit time decreased (17.5 to 9.6 h and 79.2 to 58.4 h, respectively; p < 0.05). After PA the number of fulfilled Rome criteria also decreased in group A (2.6 to 1.7; p < 0.05). In middle-aged inactive subjects with symptoms of chronic constipation, it is advisable to promote regular physical activity since it improves both the defecation pattern and rectosigmoid or total colonic transit time.


Quantitative MALDI-MS assay of steroid hormones in plasma based on hydroxylamine derivatization.
Measuring the concentrations of steroid hormones in plasma is critical for understanding their role in various vital physiological processes. The detection of underivatized steroid hormones in biofluids through mass spectrometry (MS) is typically hindered by low ionization efficiency. We described a novel matrix-assisted laser desorption/ionization-MS (MALDI-MS) approach based on hydroxylamine derivatization (HA-D) to analyze low-concentration steroid hormones in plasma. The ketonic carbonyl group containing steroid hormones could be derivatized using HA to form oxime derivatives, which considerably enhanced the MS sensitivity for detecting steroid hormones. By using the optimized conditions, estrone (E1), testosterone (T), and progesterone (Prog), could be simultaneously quantified in plasma with a limit of detection (LOD) from 0.019 to 0.031 nM, recoveries from 86% to 108%, and coefficient of variation (CV%) from 4.59% to 11.90%. HA-D/MALDI-MS exhibited higher sensitivity than those using Girard T (GT). To establish potential utility of our method, we characterized fatty liver patient plasmas to demonstrate that the HA-D/MALDI-MS procedure could generate quantitative results comparable to the current clinical liquid chromatography-electrospray ionization tandem MS (LC-ESI MS/MS) method. This approach facilitates the rapid and accurate characterization of plasma hormones, and renders the MALDI-MS approach for steroid hormones more adaptable for clinical research and use.


Simultaneous determination of 3-methoxy-4-hydroxyphenylglycol, 5-hydroxyindoleacetic acid, and homovanillic acid in cerebrospinal fluid with high-performance liquid chromatography using electrochemical detection.
An improved high-performance liquid chromatographic method with electrochemical detection (HPLC-EC) for the simultaneous determination of 3-methoxy-4-hydroxyphenylglycol (MHPG), 5-hydroxyindoleacetic acid (5-HIAA), and homovanillic acid (HVA) in cerebrospinal fluid (CSF) of humans and nonhuman primates is described. Quantitation is based on the use of an internal standard, 5-fluoro-HVA. Sample preparation consists of mixing an aliquot of CSF with a solution of the internal standard followed by ultrafiltration. The precision of the method is high, with within-run and between-run coefficients of variation of 2-6% and less than 10%, respectively, in the concentration ranges of the metabolites encountered in human lumbar CSF. Accuracy was tested by comparing the present HPLC method with specific gas chromatographic-mass spectrometric (GS-MS) assays for MHPG and HVA and a GC-MS-validated HPLC assay for 5-HIAA: the correlations obtained were 0.968 for MHPG, 0.989 for 5-HIAA, and 0.999 for HVA, with no systematic bias between the methods. The use of ascorbate as a preserving agent for monoamine metabolites in CSF was not found to be necessary when proper care was exercised in sample handling and storage. The analysis of samples with up to 2% ascorbic acid was possible as well, but MHPG had to be assayed separately using an extraction procedure and an alternative internal standard, 3-ethoxy-4-hydroxyphenylglycol.


The HB40-JUB1 transcriptional regulatory network controls gibberellin homeostasis in Arabidopsis.
The gibberellins (GAs) are phytohormones that play fundamental roles in almost every aspect of plant growth and development. Although GA biosynthetic and signaling pathways are well understood, the mechanisms that control GA homeostasis remain largely unclear in plants. Here, we demonstrate that the homeobox transcription factor (TF) HB40 of the HD-Zip family regulates GA content at two additive control levels in Arabidopsis thaliana. We show that HB40 expression is induced by GA and in turn reduces the levels of endogenous bioactive GAs by simultaneously reducing GA biosynthesis and increasing GA deactivation. Consistently, HB40 overexpression leads to typical GA-deficiency traits, such as small rosettes, reduced plant height, delayed flowering, and male sterility. By contrast, a loss-of-function hb40 mutation enhances GA-controlled growth. Genome-wide RNA sequencing combined with molecular-genetic analyses revealed that HB40 directly activates the transcription of JUNGBRUNNEN1 (JUB1), a key TF that represses growth by suppressing GA biosynthesis and signaling. HB40 also activates genes encoding GA 2-oxidases (GA2oxs), which are major GA-catabolic enzymes. The effect of HB40 on plant growth is ultimately mediated through the induction of nuclear growth-repressing DELLA proteins. Collectively, our results reveal the important role of the HB40-JUB1 regulatory network in controlling GA homeostasis during plant growth.


The effect of propolis administration on fetal development.
Propolis is one of the bee products that widely used in health therapy. However, there has no study evaluating the developmental toxicity of propolis. This study was aimed to analyze the effect of propolis administration during pregnancy on fetal development. The pregnant mice were divided into five groups including control group (Tween 80 1%), low-dose (380 mg/kg b.wt.) and high-dose (1400 mg/kg b.wt.) of water extract of propolis from Banten (WEB), and low-dose (380 mg/kg b.wt.) and high-dose (1400 mg/kg b.wt.) of ethanol extract of propolis from South Sulawesi (EES). Propolis was administered for 18 days of gestation and then sacrificed to analyze the fetal development by examining external and skeletal abnormalities. The histopathological examination of placenta was also conducted. The result showed both low-dose groups did not inhibit fetal development. However, the high-dose of EES significantly reduced the weight, crown-rump of fetuses and increased the number of resorption (p < 0.05). Fetal weight was the only significantly reduced parameter of fetal growth in the highdose group of WEB (p < 0.05). The histopathological examination of placenta showed a reduction of labyrinth development in both high-dose groups. Dose of 380 mg/kg dose of Indonesian propolis is relatively safe for consumption during pregnancy.


Synthesis and evaluation of gastroprotective activity of omeprazole related benzimidazole, imidazo[4,5-b]pyridine and quinoxaline 2-substituted derivatives.
Twenty compounds possessing benzimidazole, imidazo[4,5-b]pyridine and quinoxaline structure bearing either a substituted arylmethylmercapto- or an arylmethylsulfinyl group in position 2 were prepared in order to evaluate an antiulcer and gastroprotective activity in rat pylorus ligature, in comparison with omeprazole at the dose of 50 mg/kg after i.m. administration. One third of these compounds showed a moderate activity, being about half potent as omeprazole.