Preparation and evaluation of an oral delivery system for time-dependent colon release of insulin and selected protease inhibitor and absorption enhancer compounds.
The aim of this work was to prepare and evaluate an oral dosage form intended for time-dependent colon delivery of insulin along with a selected protease inhibitor (camostat mesilate) and absorption enhancer (sodium glycocholate). A previously described release platform, which had proven potentially suitable for the protein delivery, was exploited. Insulin compatibility with the above-mentioned adjuvants was preliminarily evaluated. For this purpose, the drug and its main degradation products were assayed by HPLC in insulin powder mixtures with camostat mesilate and/or sodium glycocholate stored 12 months at 4 degrees C. No significant decrease in protein content or increase in degradation product percentages beyond Eur. Ph. 6th Ed. limits was highlighted. Moreover, calorimetric studies performed on physical and compacted binary insulin mixtures with camostat mesilate and sodium glycocholate showed that the thermal behavior of both adjuvants was unchanged. Subsequently, tablet cores with differing compositions were prepared and spray-coated with an aqueous HPMC solution in order to obtain pulsatile delivery systems. The coated units were demonstrated to concurrently release the drug and the adjuvants in a prompt and quantitative mode after consistent lag times. Based on these results, the device was proven a potential candidate for colon delivery of insulin and the selected adjuvants.


Null mutation of the murine ATP7B (Wilson disease) gene results in intracellular copper accumulation and late-onset hepatic nodular transformation.
The Atp7b protein is a copper-transporting ATPase expressed predominantly in the liver and to a lesser extent in most other tissues. Mutations in the ATP7B gene lead to Wilson disease, a copper toxicity disorder characterized by dramatic build-up of intracellular hepatic copper with subsequent hepatic and neuro-logical abnormalities. Using homologous recombination to disrupt the normal translation of ATP7B, we have generated a strain of mice that are homozygous mutants (null) for the Wilson disease gene. The ATP7B null mice display a gradual accumulation of hepatic copper that increases to a level 60-fold greater than normal by 5 months of age. An increase in copper concentration was also observed in the kidney, brain, placenta and lactating mammary glands of homo-zygous mutants, although milk from the mutant glands was copper deficient. Morphological abnormalities resembling cirrhosis developed in the majority of the livers from homozygous mutants older than 7 months of age. Progeny of the homozygous mutant females demonstrated neurological abnormalities and growth retardation characteristic of copper deficiency. Copper concentration in the livers of the newborn homozygous null mutants was decreased dramatically. In summary, inactivation of the murine ATP7B gene produces a form of cirrhotic liver disease that resembles Wilson disease in humans and the 'toxic milk' phenotype in the mouse.


Inhibitors of metallo-β-lactamases.
The β-lactams are the most successful class of antibiotic drugs but they are vulnerable to inactivation by a growing cadre of β-lactamases that now number more than a thousand variants. β-Lactamases operate by one of two general chemical mechanisms either catalyzing β-lactam ring hydrolysis via a covalent enzyme intermediate through the aegis of an active site serine residue or through a noncovalent Zn-dependent mechanism. The Ser-β-lactamases are currently dominant in the clinic and consequently, there has been great effort to identify inhibitors and to co-formulate these with β-lactam antibiotics. Four such inhibitors are approved for human clinical use and several more are in clinical trials. Metallo-β-lactamases (MBLs), on the other hand, are only now emerging as a global threat and consequently, inhibitor discovery has lagged behind their Ser counterparts. There are now several examples of MBL inhibitors that operate either in a Zn-dependent or Zn-independent mode. The Zn-dependent compounds are more prevalent and some show efficacy in animal models of infection. These compounds function by either acting as an alternate metal ligand, usually displacing a jointly held hydroxide ion shared by enzymes with two Zn2+ ions, or alternately by striping Zn from the active site. The increase in the number of candidate MBL inhibitors over recent years reflects the growing clinical challenge of pathogens expressing these enzymes that result in a carbapenem resistance phenotype. While none of these inhibitors are yet in human clinical trials, the increasing importance of these enzymes in drug failure is a strong incentive to continue identifying and characterizing such molecules.


Canal configuration in the mesio-buccal root of maxillary first molar teeth of a Jordanian population.
The purpose of this study was to investigate the canal configuration in the mesio-buccal root of maxillary first molar teeth of a Jordanian population using a clearing technique. One hundred extracted maxillary first molars were collected from Jordanian patients. After a standard access preparation, canals were stained, decalcified, dehydrated, and then rendered clear. The prevalence of a second canal in the mesio-buccal root was 77.32%. Types IV and II canal systems were the most common types with prevalence of 35.05% and 27.83%, respectively. Additionally, 28.86% showed lateral canals mostly located in the apical third and 37.11% had intercanal communications, mainly in the middle third of the root. It was concluded that the prevalence of mesio-palatal canals was high and Types IV and II canal systems were the most prevalent.


Preferences for involvement in medical decision making: comparing British and German views.
Despite beneficial outcomes of patients' involvement in medical decision processes, it has been claimed that patients are generally not interested in medical decision making (MDM). Whereas current research focuses on actual MDM, this research explored the impact of nationality and perceptual processes of MDM. Preferences for involvement in decision making were examined in a sample of 204 German and 143 British university students. Hierarchical multiple regressions were employed to explore the link between socio-demographic information, individuals' perceived relationship with their GP, Health Locus of Control - Powerful Others, perceptions regarding the frequency and sufficiency of information provision and involvement in MDM, and individuals' preferred level of involvement and information. A significant amount of the variance in individuals preferences for involvement could be explained (Adjusted R2 = .59, p < .001). Independent t-test analyses showed that British and German perceptions of care differed significantly on a variety of different measures. Separate analyses for the German and British group highlighted cross-national differences in care and preferences for involvement. The study suggests that preferences to become involved might depend more on perceptual processes than actual involvement in decision making, and that communication and national health policy could play an important role.


Incidence of non-Hodgkin's lymphoma among individuals with chronic hepatitis B virus infection.
Although hepatitis C virus (HCV) infection has been shown to be associated with development of non-Hodgkin's lymphoma (NHL), few studies have investigated the association between chronic HBV infection and NHL. The purpose of this study was to compare the incidence of NHL between patients with and without chronic hepatitis B virus (HBV) infection. Using automated laboratory result and clinical data from two United States health systems, we identified individuals with chronic HBV infection from January 1, 1995 through December 31, 2001. Using each health system's population-based tumor registry, we identified all cases of NHL diagnosed through December 31, 2002. We excluded any individual with a history of NHL or human immunodeficiency virus (HIV). We fit Cox proportional hazards models to calculate hazard ratios comparing the incidence of NHL between chronic HBV-infected patients (N = 3,888) and patients without HBV (N = 205,203) drawn from the source populations. We identified 8 NHL cases in the chronic HBV infection cohort and 111 cases in the comparison cohort. Patients with chronic HBV infection were 2.8 times more likely to develop NHL than matched comparison patients (adjusted hazard ratio = 2.80, 95% confidence interval = 1.16-6.75), after controlling for age, race, sex, income, Charlson comorbidity index, study site, and HCV infection. chronic HBV-infected patients were nearly 3 times more likely to develop NHL than comparison patients.


Application of an improved procedure for testing the linearity of analytical methods to pharmaceutical analysis.
Examination of the requirements of the Food and Drug Administration (FDA) for evaluating the linearity of an analytical method reveals them to be unsatisfactory, in both the definition of linearity and in the specifications for testing this property of an analytical method. A new definition for linearity is proposed, along with a new method for evaluating this property of an analytical method, one that is consistent with the definition. The method is tested by re-evaluating the linearity of data collected from a Near-Infrared method of analysis of pharmaceutical preparations.


The effect of suction drains after total hip replacement.
A prospective study of 120 consecutive total hip replacements showed that deep suction drains produced maximal drainage volumes in the first 24 hours. Their continued presence resulted in minimal further drainage, did not reduce the likelihood of haematoma formation and led in some cases to the spread of skin organisms into the wound.


A comparison of 3 generic health status questionnaires among stroke patients.
Several generic multidimensional health status questionnaires are available, but it is not clear whether or not these measures are interchangeable in terms of content. Consequently, we investigated the content validity of the Sickness Impact Profile 68 (SIP68), Medical Outcome Study Short Form 36 (SF-36), and the Darmouth COOP Functional Health Assessment Charts/World Organization of National Colleges, Academies and Academic Associations of General Practitioners/Family Physicians (COOP/WONCA) charts. A total of 198 stroke patients referred to inpatient rehabilitation were interviewed 1 year poststroke. Subscales of the generic questionnaires were compared with one another and also with 3 domain-specific scales: the Barthel Index (BI) for physical functioning, the Center of Epidemiology Studies Depression (CES-D) scale for mental functioning, and the Frenchay Activities Index (FAI) for social functioning. Nonparametric Spearman correlations of at least 0.6 were accepted as evidence of content validity. Half of the physical subscales correlated with each other by at least 0.6. None of the 3 correlations in the mental domain and only 1 of 21 correlations in the social domain met the 0.6 threshold. Physical-oriented subscales correlated with BI almost as expected. In the mental domain, only the correlation of the CES-D with the SF-36 mental health scale was above 0.6, and in the social domain, only the correlation of the FAI with the SIP68 mobility range was above 0.6. The findings of this study suggest that generic health questionnaires are not mutually interchangeable. Physical and social health status of stroke patients is adequately covered by the SIP68. However, the SF-36 is more suitable for measuring the mental consequences of stroke.


Acanthamoeba infection presenting as skin lesions in patients with the acquired immunodeficiency syndrome.
Acanthamoeba organisms are a well-known, although rare, cause of central nervous system infection in immunodeficient hosts, including those with the acquired immunodeficiency syndrome. Extracerebral acanthamebiasis, with the exception of contact lens-associated keratitis, is reported but little emphasized in the literature. We describe two patients with the acquired immunodeficiency syndrome in whom skin lesions were the primary manifestations of Acanthamoeba infection. Central nervous system disease was proved in one patient and suspected, but unproved, in the other. The skin lesions exhibited an intact epidermis with suppurative inflammation of the subcutis, associated with numerous amebic cysts and trophozoites. The amebic cyst walls stained with periodic acid-Schiff and Gomori's methenamine-silver stains, creating confusion with Blastomyces dermatitidis yeast in one instance. Immunofluorescence studies and culture identified the organisms as an Acanthamoeba species. Preliminary studies in one of the cases suggested a previously undescribed Acanthamoeba species as the etiologic agent. Our experience emphasizes that skin lesions may be the presenting sign of disseminated Acanthamoeba infection in patients with the acquired immunodeficiency syndrome.


Evaluation of the Performance of Algorithms That Use Serial Hepatitis C RNA Tests to Predict Treatment Initiation and Sustained Virological Response Among Patients Infected With Hepatitis C Virus.
The structure of electronic medical record data prevents easy population-level monitoring of hepatitis C virus (HCV) treatment uptake and cure. Using an HCV registry from a public hospital system in Atlanta, Georgia, we developed multiple algorithms that use serial HCV RNA test results as proxy measures for initiation of direct-acting antiviral (DAA) treatment and sustained virological response (SVR). We calculated sensitivity and positive predictive values (PPVs) by comparing the algorithms with the DAA initiation and SVR results from the registry. From December 2013 to August 2016, 1,807 persons actively infected with HCV were identified in the registry. Of those, 698 initiated DAA treatment on the basis of medical record abstraction; of 442 patients with treatment start and/or end dates, 314 had documented SVR. Treatment algorithm 2 (a detectable HCV RNA result followed by 2 sequential HCV RNA test results) and treatment algorithm 5 (a detectable HCV RNA result followed by 2 sequential HCV RNA test results >6 weeks apart) had the highest sensitivity (87% and 85%, respectively) and PPV (80% and 82%, respectively) combinations. Four SVR algorithms relied on fulfilling treatment algorithm definitions and having an undetectable HCV RNA test result ≥12 weeks after the last HCV RNA result; sensitivity for all 4 algorithms was 79%, and PPV was 92%-93%. Algorithms using serial quantitative HCV RNA results can serve as proxy measures for evaluating population-level DAA treatment and SVR outcomes.


Central nervous involvement is common in PGM1-CDG.
PGM1, the enzyme responsible for the reversible inter-conversion of glucose-1-P and glucose-6-P, is also involved in glycosylation, leading to a wide range of clinical manifestations, such as congenital malformations, hypoglycemia, hormonal dysregulation, myopathy, hepatopathy, and cardiomyopathy. So far, PGM1 deficiency has not been associated with central nervous system involvement or intellectual disability. Seizures and neurologic involvement in PGM1-CDG were thought to be a consequence of hypoglycemia. We reviewed all reported PGM1 deficient patients for the presence of the central nervous system involvement, their treatment and disease history. We detected 17 patients out of the 41 reported PGM1-CDG cases with significant neurologic involvement. Several of these patients had no severe hypoglycemic episodes, or were adequately treated for hypoglycemia with no recurrent episodes of low blood sugars, while one patient had no reported hypoglycemic episodes. We suggest that neurological symptoms are frequent in PGM1-CDG and could present even in the absence of hypoglycemia. The central nervous system should be assessed early on during the diagnostic process to optimize outcome in patients with PGM1-CDG.


Steroidal control of rat uterine 17 beta-hydroxysteroid dehydrogenase activity.
The interconversion of estradiol-17 beta and estrone in the rat uterus is due to the action of 17 beta-hydroxysteroid dehydrogenase. Whole uteri or 800 x g supernatant fractions of the uteri were incubated in the presence of [3H] estradiol-17 beta and NAD at 37 degrees C for 3 h or 1 h, respectively. In the mature rat uterus the oxidation of estradiol-17 beta and estrone was dependent on the stage of the estrous cycle, suggesting hormonal control. The 17 beta-hydroxysteroid dehydrogenase activity was highest at estrus (200 fmol estrone) and lowest at diestrus (80 fmol estrone). An enhancement of activity occurred when adult rats at each stage of the estrous cycle were administered estradiol-17 beta, while progesterone administration at each stage resulted in decreased enzyme activity. The uterine 17 beta-hydroxysteroid dehydrogenase activity of estradiol-17 beta treated ovariectomized rats was time and dose dependent but decreased when progesterone was administered with or without estradiol-17 beta administration. These results suggest that estradiol-17 beta caused an increase in enzyme activity that was inhibitable by progesterone in the rat uterus. The increased 17 beta-hydroxysteroid dehydrogenase activity may reflect a specific response of the rat uterus to estradiol-17 beta.


Synthesis, SAR study, and biological evaluation of novel quinoline derivatives as phosphodiesterase 10A inhibitors with reduced CYP3A4 inhibition.
A novel class of phosphodiesterase 10A inhibitors with potent PDE10A inhibitory activity and reduced CYP3A4 inhibition was designed and synthesized starting from 2-[4-({[1-methyl-4-(pyridin-4-yl)-1H-pyrazol-3-yl]oxy}methyl)phenyl]quinoline (1). Replacement of pyridine ring of 1 with N-methyl pyridone ring drastically improved CYP3A4 inhibition, and further optimization of these quinoline analogues identified 1-methyl-5-(1-methyl-3-{[4-(quinolin-2-yl)phenoxy]methyl}-1H-pyrazol-4-yl)pyridin-2(1H)-one (42b), which showed potent PDE10A inhibitory activity and a good CYP3A4 inhibition profile. A PET study with (11)C-labeled 42b indicated that 42b exhibited good brain penetration and specifically accumulated in the rodent striatum. Further, oral administration of 42b dose-dependently attenuated phencyclidine-induced hyperlocomotion in mice with an ED50 value of 2.0mg/kg and improved visual-recognition memory impairment at 0.1 and 0.3mg/kg in mice novel object recognition test.


Rapid prediction of steady-state serum theophylline concentration in patients treated with intravenous aminophylline.
In 15 acutely ill asthmatics the steady-state serum theophylline concentration was predicted by the method of Chiou et al. using two serum concentration measurements obtained 1 and 5 h after starting a continuous infusion of aminophylline. Two theophylline assays with different precision characteristics were compared. With a precise HPLC-assay the prediction was excellent: prediction error (predicted minus measured concentration) = -0.22 +/- 1.97 mg/l (mean +/- SD); r = 0.922. When the theophylline concentration was determined by a rapid enzyme immunoassay of lower precision, but convenient for clinical use, the prediction was less accurate (prediction error = 0.58 +/- 3.88, r = 0.852). However, it was still clearly superior to dosing recommendations based on the population average of theophylline clearance, even after taking into consideration the effect of smoking, congestive heart failure and cirrhosis (prediction error = 3.62 +/- 13.36, r = 0.560). As employed in this study, the method may be useful in helping the physician to choose the optimal dose in severely ill asthmatics.


Neuropeptidergic innervation of intramuscular hemangiomas.
Intramuscular hemangiomas are idiopathic lesions which are either tumoral or developmental in origin. A close association of abnormal blood vessels with nerve fibers is found and may suggest that nerves have a primary inciting role in the development of these lesions. In the current study, the number of nerve fibers in different zones around the tumors, as well as the type of neuropeptides present in these fibers, was quantitatively assessed by computer-assisted image analysis of immunohistochemical staining of histological slides. The number of nerve fibers as determined by positive staining by anti-protein S-100 antibodies was found to be elevated in the immediate vicinity of the abnormal blood vessels. The density of the nerve fibers rapidly declined with increasing distance from the hemangiomas, reaching normal values at distances of over 2 mm. Furthermore, hemangiomas contain a significantly higher number of calcitonin gene-related peptide (CGRP), substance P, and Met-enkephalin-positive fibers. The most significant rise in number is that of CGRP-positive fibers. This neuropeptide is a known mitogen, which could be responsible for the growth of the hemangiomatous blood vessels. Substance P is a nociceptive neurotransmitter and its presence can explain the pain which often accompanies even tiny intramuscular hemangiomas.


Regulation of MHC class I heterodimer stability and interaction with TAP by tapasin.
Major histocompatibility complex (MHC) class I molecules are heterodimers of a class I heavy chain and beta 2-microglobulin that bind peptides supplied by the MHC region-encoded transporters associated with antigen processing (TAP). Peptide binding by class I heterodimers is necessary for their maturation into stable complexes and is dependent on their physical association with TAP. In human mutant 721.220 cells, however, a novel genetic defect causes the failure of class I heterodimers to associate with TAP. This deficiency correlates with lack of expression of a glycoprotein, tapasin (TAP-associated glycoprotein), which has been found in association with class I heterodimers and TAP. Employing a transcomplementation analysis, we obtained evidence co-localizing the genetic defect of mutant 220 cells and the structural or a regulatory gene controlling the expression of tapasin on the short arm of chromosome 6, which includes the MHC. Expression of tapasin and the normal interaction of class I heterodimers with TAP are concomitantly restored, indicating the probable function of tapasin as a physical link between these complexes. In further support of this model, the absence of tapasin in mutant 220 cells correlates with reduced class I heterodimer stability, suggesting that tapasin may stabilize class I heterodimers and thereby enhance their association with TAP. These results further implicate tapasin in a mechanism that promotes peptide binding by class I heterodimers through their interaction with TAP.


Positron emission tomography using [18F]Galacto-RGD identifies the level of integrin alpha(v)beta3 expression in man.
The integrin alpha(v)beta3 plays a key role in angiogenesis and tumor cell metastasis and is therefore an important target for new therapeutic and diagnostic strategies. We have developed [18F]Galacto-RGD, a highly alpha(v)beta3-selective tracer for positron emission tomography (PET). Here, we show, in man, that the intensity of [18F]Galacto-RGD uptake correlates with alpha(v)beta3 expression. Nineteen patients with solid tumors (musculoskeletal system, n = 10; melanoma, n = 4; head and neck cancer, n = 2; glioblastoma, n = 2; and breast cancer, n = 1) were examined with PET using [18F]Galacto-RGD before surgical removal of the tumor lesions. Snap-frozen specimens (n = 26) were collected from representative areas with low and intense standardized uptake values (SUV) of [18F]Galacto-RGD. Immunohistochemistry was done using the alpha(v)beta3-specific antibody LM609. Intensity of staining (graded on a four-point scale) and the microvessel density of alpha(v)beta3-positive vessels were determined and correlated with SUV and tumor/blood ratios (T/B). Two tumors showed no tracer uptake (mean SUV, 0.5 +/- 0.1). All other tumors showed tracer accumulation with SUVs ranging from 1.2 to 10.0 (mean, 3.8 +/- 2.3; T/B, 3.4 +/- 2.2; tumor/muscle ratio, 7.7 +/- 5.4). The correlation of SUV and T/B with the intensity of immunohistochemical staining (Spearman's r = 0.92; P < 0.0001) as well as with the microvessel density (Spearman's r = 0.84; P < 0.0001) were significant. Immunohistochemistry confirmed lack of alpha(v)beta3 expression in normal tissue (benign lymph nodes, muscle) and in the two tumors without tracer uptake. Molecular imaging of alpha(v)beta3 expression with [18F]Galacto-RGD in humans correlates with alpha(v)beta3 expression as determined by immunohistochemistry. PET with [18F]Galacto-RGD might therefore be used as a new marker of angiogenesis and for individualized planning of therapeutic strategies with alpha(v)beta3-targeted drugs.


Ancestral polymorphisms in Drosophila pseudoobscura and Drosophila miranda.
Ancestral polymorphisms are defined as variants that arose by mutation prior to the speciation event that generated the species in which they segregate. Their presence may complicate the interpretation of molecular data and lead to incorrect phylogenetic inferences. They may also be used to identify regions of the genome that are under balancing selection. It is thus important to take into account the contribution of ancestral polymorphisms to variability within species and divergence between species. Here, we extend and improve a method for estimation of the proportion of ancestral polymorphisms within a species, and apply it to a dataset of 33 X-linked and 34 autosomal protein-coding genes for which sequence polymorphism data are available in both Drosophila pseudoobscura and Drosophila miranda, using Drosophila affinis as an outgroup. We show that a substantial proportion of both X-linked and autosomal synonymous variants in these two species are ancestral, and that a small number of additional genes with unusually high sequence diversity seem to have an excess of ancestral polymorphisms, suggestive of balancing selection.


Immediate shock deficit in fear conditioning: effects of shock manipulations.
Pavlovian contextual fear conditioning occurs when an aversive unconditional stimulus (US), such as a footshock, is presented to a rat shortly after it is placed in an experimental context. Contextual fear conditioning does not occur when the shock is presented immediately upon placement of the rat in the novel chamber. In the present study, the authors report that increasing either the number of immediate shock sessions (Experiment 1) or the immediate shock duration (Experiment 2) did not reverse this deficit. However, immediate shock seems to sensitize subsequent context conditioning (Experiment 3). These findings suggest that the associative deficit produced by immediate shock is not related to the rat's ability to process the footshock US.


Naringin protects against bone loss in steroid-treated inflammatory bowel disease in a rat model.
We observed the effects of naringin on bone loss in glucocorticoid-treated inflammatory bowel disease (IBD) in a rat model. The IBD model was established in Sprague-Dawley rats by administering 5.0% dextran sodium sulfate. Dexamethasone (DEX) and naringin were given at the second week. Blood, colon and bone samples were collected for biomarker assay, histological analysis or microCT analysis. Superoxide dismutase, catalase and malonaldehyde were measured in bone. A significant decrease of procollagen type 1 N-terminal propeptide (P1NP) level was observed in DEX-treated IBD groups compared with the control (p < 0.05). P1NP levels were dose-dependently increased in the presence of naringin intervention. Bone loss and decreased bone biomechanical properties were observed in DEX-treated IBD rats compared with control rats (p < 0.01). Naringin intervention protected against bone loss and decreased bone biomechanical properties. Bone formation related gene mRNA expressions were significantly decreased in DEX-treated IBD rats compared with control rats. Naringin administration reversed the down-regulation of the expressions of those genes. Naringin treatment reduced the oxidative stress in bone from DEX-treated IBD rats. Our data indicated that naringin may have great potential for the treatment of bone loss in glucocorticoid-treated IBD patients via blocking oxidative stress and promoting bone formation.


Role of aggregates in Claisen acylation reactions of imidazole, pyrazole, and thioesters with lithium enolates in THF.
[formula: see text] Although phenyl esters react with both monomers and dimers or tetramers of two lithium enolates in THF, the reactions of phenyl thiobenzoates are relatively much faster with the monomers. Similarly, imidazole esters react primarily with the monomers but pyrazole esters react with monomers and aggregates. The results are rationalized by a mechanism in which coordination with two lithium cations within an enolate aggregate is required for the reaction of aggregates to compete with monomers.


Depressed neutrophil motility in patients with recurrent herpes simplex virus infections: in vitro restoration with levamisole.
The random migration and chemotaxis of neutrophils from 10 patients with recurrent infections with herpes simplex virus were assessed. Chemotaxis to endotoxin-activated serum was markedly diminished in this group, but random migration was only mildly reduced, and chemotaxis to hydrolyzed casein was normal. The effects of the immunostimulant drug levamisole on the chemotaxis of neutrophils were assessed both in vitro and in patients after 150 mg of the drug was taken daily for three days. The defective chemotactic response could be corrected in vitro by treatment of the neutrophils with 10(-3) M levamisole. Of 10 patients treated with levamisole, five showed improved chemotaxis after three days, but no improvement in neutrophil function was observed in the other patients.


In vitro metabolism of human and salmon calcitonins in rat liver and kidney evaluated by liquid chromatography-tandem mass spectrometry.
1. Using LC-MS and LC-MS/MS, an in vitro study was conducted on the metabolism of human calcitonin (hCT) and salmon calcitonin (sCT) in rat liver and kidney to determine the rates of metabolism and the positions of hydrolytic cleavage in both peptides. 2. In lysosomal fractions of rat liver and kidney, hCT was degraded 9-12 times faster than sCT. Many metabolites of hCT were produced in the lysosomal fractions, whereas the metabolites of sCT were scarcely found. 3. In the case of the cytosolic fractions, three positions of initial endoproteolytic cleavage were found in hCT, leading to the production of many peptide fragments via subsequent exoproteolytic metabolism. The initial cleavage position of sCT could not be identified precisely, but it was postulated that the rate-determining step in the metabolism of sCT is the endoproteolytic hydrolysis. 4. The studies using pure proteases and protease inhibitors indicated that the metabolism of calcitonins proceeds by initial endoproteolytic cleavage and subsequent exoproteolytic digestion, catalysed by an aspartate-protease in lysosomes and by a metalloprotease and cysteine-protease in combination in the cytosol. 5. The result suggested that the higher in vivo pharmacological activity of sCT compared with that of hCT may be due to a slower metabolism of the former.


Splicing-dependent and -independent modes of assembly for intron-encoded box C/D snoRNPs in mammalian cells.
In mammalian cells, all small nucleolar RNAs (snoRNAs) that guide rRNA modification are encoded within the introns of host genes. An optimal position about 70 nts upstream of the 3' splice site of the host intron is critical for efficient expression of box C/D snoRNAs in vivo, suggesting synergy with splicing. Here, we have used a coupled in vitro splicing-snoRNA processing system to demonstrate that assembly of box C/D snoRNP proteins is the step affected by snoRNA location, and that active splicing is essential for snoRNP assembly. Splicing blockage experiments further reveal that snoRNP proteins bind specifically at the spliceosomal C1 complex stage. In contrast, splicing-independent snoRNP assembly can occur in vitro on snoRNAs that possess stable external stems. In vivo analyses confirm that a stable stem can compensate for the unusual position of those few box C/D snoRNAs located far from the 3' splice site of their host intron.


Gene structure and expression of a novel Euglena gracilis chloroplast operon encoding cytochrome b6 and the beta and epsilon subunits of the H(+)-ATP synthase complex.
The genes encoding cytochrome b6 of the chloroplast cytochrome b6/f complex (petB) and the ATP synthase CF1-beta subunit (atpB) and epsilon-subunit (atpE) were identified on the EcoD fragment of the Euglena gracilis chloroplast genome. The complete nucleotide sequence of these three genes was determined. The petB-atpB-atpE genes are cotranscribed as a tricistronic operon. This gene organization differs from that of land plants in which atpB-atpE form a discistronic operon, and petB is within the psbB-ycf8-psbH-petB-petD operon. Euglena cytochrome b6 and the beta-subunit of the chloroplast ATP synthase are very similar in derived amino acid sequence to the corresponding gene products from other organisms. The epsilon-subunit of the chloroplast ATP synthase complex is more divergent. In Euglena, the petB-atpB-atpE genes contain introns, including two twintrons, at eight different positions. All of the intron positions were confirmed by analysis of cDNAs. Two independent intercistronic RNA processing events and 11 splicing reactions lead to the accumulation of the mature petB, atpB and atpE monocistronic mRNAs.


Isolation from bovine brain and structural characterization of HBNF, a heparin-binding neurotrophic factor.
A heparin-binding protein with neurotrophic activity for perinatal rat neurons, termed HBNF, was purified to homogeneity from bovine brain utilizing pH 4.5 extraction, ammonium sulfate precipitation, cation exchange and heparin-Sepharose affinity chromatographies, and reverse phase HPLC. In the presence of protease inhibitors during extraction, a protein with an apparent molecular weight of 18 kDa was obtained in a yield of approximately 0.5 mg/kg brain tissue. The amino acid sequence of the first 114 residues of HBNF was determined and found to highly homologous to the cDNA-derived amino acid sequence of human HBNF, a 136-residue protein. Bovine and human HBNFs have identical molecular weights as judged by SDS gel electrophoresis and very similar amino acid compositions. This and overall sequence conservation suggest that bovine HBNF is also a 136 amino acid protein with a calculated molecular weight of approximately 15.5 kDa. The apparent discrepancy between calculated and observed molecular weights of bovine HBNF (and of human HBNF of which the complete sequence is known) is most likely a result of the highly basic nature of HBNF. If protease inhibitors were omitted during tissue extraction, two additional proteins with lower apparent molecular weights and identical N-terminal sequences were isolated, with the smallest forms being the major product. Amino acid analysis showed that the smaller forms correspond to C-terminally truncated HBNFs with calculated molecular weights of 13.6 and 12.4 kDa, lacking approximately 14 and 22 residues. Comparison of the HBNF protein sequence with sequences stored in the Protein Identification Resource/Genbank databases reveals high homology to the translation product of the MK-1 gene, which is retinoic acid-inducible in embryonic carcinoma cells and developmentally expressed during gestation in mice.


Early diagnosis of glaucoma. II. The value of the initial examination in ocular hypertension.
In a group of 46 patients with moderately elevated intraocular pressure (22-30 mmHg), signs of early glaucoma were found in half of them after a follow-up of 5-7.5 years. Most of the patients showed only a small increase in optic disc cupping revealed by stereophotography. Initial IOP higher than 25 mmHg, large pressure differences between the two eyes, large diurnal pressure variation, occurrence of pseudoexfoliation, pressure drop of more than 8 mmHg after topical epinephrine, and a pressure rise of more than 4 mmHg after water loading were more frequent in the group of patients who developed signs of glaucoma. Age, a positive family history, occurrence of cardiovascular disease, and the cup/disc ratio did not differ in the two groups of patients. The pilocarpine test and steroid provocation had no predictive value. Surprisingly, there was a small drop of pressure after topical steroids in 6 of the 8 patients with pseudoexfoliation.


Association between the IL7R T244I polymorphism and multiple sclerosis: a meta-analysis.
Previously published analyses of the association between the interleukin 7 receptor (IL7R) T244I polymorphism (rs6897932) and multiple sclerosis (MS) have yielded conflicting results. We performed a meta-analysis to assess whether the combined data showed this association, and to investigate its effect size. We analyzed 10 studies identified from PubMed (12,185 MS patients and 15,855 controls) and calculated the odds ratios (ORs) and 95% confidence intervals (CIs) for the C-allele, the C/C genotype (recessive effect) and the C/C + C/T (dominant effect) genotype. Heterogeneity within and between studies was observed: allele C: Q = 30.86, P = 0.002; genotype C/C: Q = 30.28, P = 0.003. Using a random-effects model, the C-allele and the C/C genotype were associated with MS (OR = 1.11, 95% CI = 1.04-1.19, P = 0.001 for the C-allele; OR = 1.15, 95% CI = 1.06-1.24, P = 0.0009 for the C/C genotype). The C/C + C/T genotype was also associated with MS using a fixed-effects model (OR = 1.15, 95% CI = 1.05-1.26, P = 0.003). There was no significant publication bias among the selected studies according to the funnel plot. We also performed the analysis on a European subgroup. This revealed an association between IL7R T244I and MS (P < 0.00001 for the C-allele and the C/C genotype; P = 0.0004 for the C/C + C/T genotype), no heterogeneity was observed (allele C: P = 0.07; genotype C/C: P = 0.10). In conclusion, the meta-analysis demonstrated that the IL7R T244I polymorphism was associated with susceptibility to MS.


Sensory gating of auditory evoked potentials in rats: effects of repetitive stimulation and the interstimulus interval.
In the P50 gating or conditioning-testing (C-T) paradigm, the P50 response, a small positive midlatency ( approximately 50 ms after stimulus onset) component of the human auditory evoked potential (AEP), is reduced towards the second click (S2) as compared to the response to the first click (S1). This phenomenon is called sensory gating. The putative function of sensory gating is thought to protect subjects from being flooded by irrelevant stimuli. Comparative studies have been done in rats in order to elucidate the underlying neural substrate of sensory gating. However, for a direct comparison of rat and human AEP components, it is imperative for both components to show similar characteristics. The amount of sensory gating in humans is dependent on repetitive stimulation and the interstimulus interval (ISI). In the present study effects of repetitive stimulation (Experiment 1) and various ISIs (Experiment 2) were determined on rat AEP components. The results demonstrate that gating is not limited to a restricted cortical area or a single midlatency component and that repetitive stimulation and ISI affect gating of several rat AEP components. Components such as the vertex P17 and N22 show a decrease in gating within several S1-S2 presentations, mainly due to a decrease in amplitude to S1 (Experiment 1). Gating for vertex components (such as the P17, N22 and N50) is ISI dependent (Experiment 2), but there is no interval in the 200-600 ms range at which optimal gating occurs. The ISI effects on gating are due to an increase of the amplitude to S2. The results have implications for the discussion about the rat homologue of the human P50.


Combined bead-based multiplex detection of RNA and protein biomarkers: Implications for understanding the time course of skeletal muscle injury and repair.
Biological response to skeletal muscle injury time course is generally classified as initial (elevated within first 4-h), delayed (elevated at 24-h), and/or prolonged (elevated at 4-h and sustained to 24-h). Accurate description of this process requires the ability to measure a robust set of RNA and protein biomarkers, yet such an approach is not common and not always feasible. This method proposes a novel experimental approach that focuses on the use of bead-based multiplex detection to measure mRNA, lncRNA, cytokines, soluble cytokine receptors, and myokines at 4-h and 24-h post muscle injury. We used an extreme aerobic exercise session (half-marathon race) to create a consistent muscle injury stimulus via oxidative stress and eccentric contractions. Venous blood samples were analyzed to determine the change in 90 targets. Specifically, we identified 14 mRNA, 2 lncRNA, 4 cytokines, and 5 myokines that had only an initial response (change at 4-h). We identified 2 mRNA, 2 cytokines, 13 soluble cytokine receptors, and 1 myokine that had only a delayed response (change at 24-h). Finally, we identified 18 mRNA, 4 lncRNA, 6 myokines and 15 cytokines that had a prolonged response (change at 4-h and sustained at 24-h). We found 4 targets to be undetectable or having no response relative to muscle injury recovery. These findings demonstrate the interplay between RNA and protein biomarkers in response to skeletal muscle injury. This novel experimental application of bead-based multiplexing is applicable to a variety of clinical models that involve muscle injury and/or wasting.


Effects of morphine dependence and withdrawal on levels of neurosteroids in rat brain.
To investigate the effects of morphine dependence and withdrawal on the concentrations of neurosteroids in rat brain. A method of simultaneous quantification of neurosteroids by gas chromatography-mass spectrometry (GC-MS) had been established. The chronic morphine administration (ip) resulted in a marked decrease in the brain concentrations of pregnenolone (PREG), progesterone (PROG), and pregenenolone sulfate (PREGS) in rats killed 6 h after the last treatment. In contrast, there were no significant effects of morphine dependence on the brain concentrations of allopregnanolone (AP), dihydroepiandrosterone (DHEA), and dihydroepiandrosterone sulfate (DHEAS). Naloxone-induced withdrawal produced a significant increase in the concentrations of PREG, PROG, AP, DHEA, PREGS, and DHEAS as compared with the control group. Morphine dependence and withdrawal affected the concentrations of neurosteroids in rat brain, which suggests that endogenous neurosteroids in brain might be related to the development of morphine dependence and withdrawal.


Imaging the female pelvis at 3.0 T.
Three-Tesla whole body imaging is rapidly becoming part of routine clinical practice. Although it is generally thought that pelvic imaging at 3.0 T will be beneficial because of increased signal to noise and greater spectral separation, adjustments in protocol and sequence parameters are necessary to optimize image quality. The question remains as to whether 3.0-T imaging will offer further benefits beyond 1.5 T in terms of lesion characterization and functional imaging. This article aims to address safety concerns and to illustrate the potential benefits and technical challenges of imaging the female pelvis at 3.0 T. Imaging protocols and sequence parameters for routine gynecologic indications are suggested, and potential clinical applications at 3.0 T are discussed such as magnetic resonance spectroscopy, perfusion, diffusion weighted imaging, and the use of alternate contrast agents.


Evaluation of carboxymethylpullulan as a novel carrier for targeting immune tissues.
To demonstrate the potential of carboxymethylpullulan (CMPul) as a carrier for targeting immune tissues, and to find whether immune tissues could be set as the target of an immunosuppressant to treat autoimmune diseases. The biodistribution of CMPul was investigated to evaluate its potency as a carrier for targeting immune tissues. Furthermore, an immunosuppressant-CMPul conjugate was prepared and its suppressive effect on rat adjuvant arthritis was examined. The disappearance rate of 3H-labeled CMPul from the blood circulation was much slower than that of 3H-labeled pullulan (Pul) after intravenous injection to normal rats. The concentration of 3H-labeled CMPul in the spleen and lymph nodes was much higher than that of 3H-labeled Pul at 24 hours after the injection, whereas the concentration of 3H-labeled CMPul in the liver was significantly lower than that of 3H-labeled Pul. A similar targeting property of 3H-labeled CMPul for these immune tissues was observed in arthritic rats. A conjugate composed of a novel immunosuppressant PA-48153C and CMPul showed a suppressive effect on rat adjuvant arthritis judging from a reduction of the arthritic index and spleen weight and an increase of body weight. CMPul is expected to be a promising carrier for targeting immune tissues with an immunosuppressant to enable treatment of autoimmune diseases.


Validation of an immunoassay for soluble Klotho protein: decreased levels in diabetes and increased levels in chronic kidney disease.
The Klotho gene has been identified as an aging suppressor gene that encodes a transmembrane protein, which is expressed primarily in renal tubules. There are 2 forms of Klotho, membrane and secreted. However, there is a paucity of data on levels of soluble Klotho in diseases like diabetes and kidney disease. We validated an enzyme-linked immunosorbent assay for Klotho and quantitated Klotho levels separately in patients with diabetes and also in patients with chronic kidney disease (CKD). The Klotho assay showed good precision and was linear down to 19 ng/mL. There were no significant effects on Klotho levels with the addition of common interferents such as ascorbate, triglycerides, or hemolysis; only bilirubin (250 mg/L) significantly reduced Klotho levels (P < .05). There was a significant reduction in Klotho levels in samples with glycated hemoglobin (HbA(1c)) levels of 6.5% or more compared with control samples (HbA(1c) < 6.5%; P < .001). We also documented significantly higher levels of Klotho with CKD. Thus, we validated an assay for Klotho and made the novel observation that levels are decreased in diabetes and increased in CKD.


The amyloid paradox: amyloid-beta-metal complexes can be neurotoxic and neuroprotective.
Senile plaques in the brains of people with Alzheimer disease (AD) are primarily composed of the amyloid-beta (Abeta) peptide and contain substantially elevated levels of iron, copper and zinc. These metals bind to Abeta and have been reported to increase the toxicity of Abeta to cultured neurones. Other reports have demonstrated that Abeta can reduce the neurotoxicity of metal ions, suggesting that the interaction can, under some circumstances, be protective. To investigate these apparently conflicting results, human Abeta1-42 was co-injected with iron, copper or zinc (at the concentrations found in plaques) into rat cerebral cortex, and the resulting numbers of dying neurones were compared. It was found that Abeta complexed with either iron or zinc was more toxic than Abeta alone. In contrast, Abeta-copper complexes were not neurotoxic. Surprisingly, we observed that when iron or copper were combined with Abeta, the neurotoxicity of these metals was substantially reduced, suggesting that Abeta may help to limit the toxicity of redox-active metal ions, thereby assisting the antioxidant defence of the brain. Thus paradoxical effects occur when Abeta complexes with metal ions, where Abeta-metal complexes are capable of being neurotoxic and neuroprotective.


Interactions between occupational exposure to extremely low frequency magnetic fields and chemicals for brain tumour risk in the INTEROCC study.
In absence of clear evidence regarding possible effects of occupational chemical exposures on brain tumour aetiology, it is worthwhile to explore the hypothesis that such exposures might act on brain tumour risk in interaction with occupational exposure to extremely low frequency magnetic fields (ELF). INTEROCC is a seven-country (Australia, Canada, France, Germany, Israel, New Zealand and UK), population-based, case-control study, based on the larger INTERPHONE study. Incident cases of primary glioma and meningioma were ascertained from 2000 to 2004. Job titles were coded into standard international occupational classifications and estimates of ELF and chemical exposures were assigned based on job-exposure matrices. Dichotomous indicators of cumulative ELF (≥50th vs <50th percentile, 1-4 year exposure time window) and chemical exposures (ever vs never, 5-year lag) were created. Interaction was assessed on both the additive and multiplicative scales. A total of 1939 glioma cases, 1822 meningioma cases and 5404 controls were included in the analysis, using conditional logistic regression. There was no clear evidence for interactions between ELF and any of the chemical exposures assessed for either glioma or meningioma risk. For glioma, subjects in the low ELF/metal exposed group had a lower risk than would be predicted from marginal effects. Results were similar according to different exposure time windows, to cut-points of exposure or in exposed-only analyses. There was no clear evidence for interactions between occupational ELF and chemical exposures in relation to glioma or meningioma risk observed. Further research with more refined estimates of occupational exposures is recommended.


Up-scaling expectations among Pakistan's HIV bureaucrats: entrepreneurs of the self and job precariousness post-scale-up.
Existing research has documented how the expansion of HIV programming has produced new subjectivities among the recipients of interventions. However, this paper contends that changes in politics, power and subjectivities may also be seen among the HIV bureaucracy in the decade of scale-up. One year's ethnographic fieldwork was conducted among AIDS control officials in Pakistan at a moment of rolling back a World Bank-financed Enhanced Programme. In 2003, the World Bank convinced the Musharraf regime to scale up the HIV response, offering a multimillion dollar soft loan package. I explore how the Enhanced Programme initiated government employees into a new transient work culture and turned the AIDS control programmes into a hybrid bureaucracy. However, the donor money did not last long and individuals' entrepreneurial abilities were tested in a time of crisis engendered by dependence on aid, leaving them precariously exposed to job insecurity, and undermining the continuity of AIDS prevention and treatment in the country. I do not offer a story of global 'best practices' thwarted by local 'lack of capacity', but an ethnographic critique of the transnational HIV apparatus and its neoliberal underpinning. I suggest that this Pakistan-derived analysis is more widely relevant in the post-scale-up decade.


Comparison of two models for estimation of variance components in a sample of Spanish Holstein Friesians.
Two models for estimating genetic parameters in animal populations were compared. The cheapest, in terms of computing costs, was based on a sire model and the most expensive on an animal model. In order to check the accuracy of both methods they were applied to the best available sample of Spanish Friesian cattle. Milk production of each cow, as well as fat contents were simulated using prior estimates of genetic additive, permanent environment and residual variances, keeping the actual mating scheme, so that the simulation mimicked the actual population structure. Results, under the above premises, show no advantages of the more costly procedure over the cheaper one, suggesting that a sire model based estimating procedure for genetic parameters may be preferred when a small number of individuals, little pedigree information and highly disequilibrated distribution of effects characterize the data. ZUSAMMENFASSUNG: Vergleich zweier Methoden der Varianzkomponentenschätzung beim Spanischen Holstein Zwei Modelle zur Schätzung von genetischen Parametern wurden verglichen. Das im Hinblick auf die Rechenleistung günstigere Modell war ein Vatermodell, das aufwendigere Modell ein Tiermodell. Um die Genauigkeit der Modelle in der Praxis zu prüfen, wurden sie auf verfügbares Material der spanischen Holstein-Friesian Population angewendet. Sowohl die Milchleistung jeder Kuh als auch der Fettgehalt wurden simuliert mit Hilfe von Schät- zwerten der additiv-genetischen Varianz, der permanenten und der residuellen Umweltvarianz. Dabei wurde das tatsächliche Ungleichgewicht in der Population berücksichtigt. Unter den obengenannten Voraussetzungen zeigen die Ergebnisse keinen Vorteil der rechnerisch aufwendigeren Prozedur gegenäber der einfacheren. Die Schlußfolgerung lautet, daß das Vatermodell unter den Bedingungen der kleinen Tierzahlen, wenig Pedigree-Informationen und einer stark unbalan- cierten Verteilung der Tiere auf die einzelnen Effektklassen zu bevorzugen ist.


Restoration of cytochrome P450 2C11 in vitamin A-deficient rat liver by exogenous androgen.
Down-regulation of microsomal androgen-dependent CYP2C11 is produced in male rat liver by dietary vitamin A deficiency. Decreased circulating androgen concentrations also occur in vitamin A-deficient male rats. Both effects are prevented by addition of all-trans-retinoic acid to the diet. The present study evaluated directly whether androgen deficiency may be responsible for the down-regulation of 2C11 in vitamin A-deficient male rats. The major finding was that subcutaneous administration of the androgen methyltrienolone (MT) during the final week of the study restored CYP2C11 protein and its associated steroid 16alpha-hydroxylation activities to control levels; CYP2C11 mRNA was also restored. Despite the efficient restoration of CYP2C11 at a pretranslational level, no alteration in vitamin A status was apparent and animals remained vitamin A deficient after MT treatment. The possibility was assessed that vitamin A can maintain the microsomal content of CYP2C11 in normal liver. However, in contrast to MT, administration of ATRA to gonadectomized male rats did not restore 2C11 in liver. These findings establish that the major effect of vitamin A deficiency on CYP2C11 in male rat liver is mediated indirectly by androgen deficiency.


Traditional Chinese Medicine for Cardiovascular Disease: Evidence and Potential Mechanisms.
Traditional Chinese medicine (TCM) has more than 2,000 years of history and has gained widespread clinical applications. However, the explicit role of TCM in preventing and treating cardiovascular disease remains unclear due to a lack of sound scientific evidence. Currently available randomized controlled trials on TCM are flawed, with small sample sizes and diverse outcomes, making it difficult to draw definite conclusions about the actual benefits and harms of TCM. Here, we systematically assessed the efficacy and safety of TCM for cardiovascular disease, as well as the pharmacological effects of active TCM ingredients on the cardiovascular system and potential mechanisms. Results indicate that TCM might be used as a complementary and alternative approach to the primary and secondary prevention of cardiovascular disease. However, further rigorously designed randomized controlled trials are warranted to assess the effect of TCM on long-term hard endpoints in patients with cardiovascular disease.


Effect of various types of exercise training on 5'-nucleotidase and adenosine deaminase activities in rat heart: influence of a single bout of endurance exercise.
The aim of the present study was to find out whether activities of the enzymes controlling adenosine metabolism, 5'-nucleotidase (5NT) and adenosine deaminase (ADA), in the left ventricle of the rat's heart change after 6 weeks of endurance or sprint training. Additionally, an influence of a single bout of endurance exercise till exhaustion on activities of these enzymes was investigated in sedentary and trained rats. The rats were divided into three groups: (1) sedentary controls (C), (2) endurance-trained (ET), and (3) sprint-trained (ST). It was shown that both types of training increased 5NT, but did not change ADA activity in the rat heart. Acute exercise till exhaustion did not affect 5NT activity in the heart taken from C and ST rats, but decreased its activity in the ET group. The heart ADA activity after exhaustive exercise increased in C and in ET group, but decreased in ST animals. It is concluded that physical training affects cardiac adenosine metabolism and the type of training may exert an influence on purine nucleotides metabolism in the heart during exhaustive exercise.


SW-163A and B, novel immunosuppressants produced by Streptomyces sp.
SW-163A and B, novel immunosuppressants, were isolated from the culture broth of Streptomyces sp. SNA15896. The molecular formulas of SW-163A and B were determined as C34H42N2O12 and C33H40N2O12 based on the HRFAB-MS analysis, respectively. Both compounds were elucidated to be a large ring ester structure through spectroscopic analyses including various NMR measurements. SW-163A and B showed immunosuppressive and antimicrobial activities in vitro.


Effect of angiotensin-converting enzyme gene polymorphism on pregnancy outcome, enzyme activity, and zinc concentration.
To evaluate the effect of angiotensin-converting enzyme (ACE) genotypes on pregnancy outcome, the incidence of pregnancy-induced hypertension, and changes in blood pressure (BP) during pregnancy; and the relationship between plasma ACE activities and plasma and erythrocyte zinc concentrations in each genotype. The subjects (n = 191) were selected from 580 indigent African-American pregnant women who enrolled toward the end of a trial to evaluate the effect of zinc supplementation on pregnancy outcome. This selection resulted in 93 subjects who received zinc and 98 who received placebo. Sample size was calculated with a 0.50 correlation coefficient between plasma ACE activities and zinc levels and a power of 80%. This calculation indicated that the sample size in each ACE genotype should be more than 28. Angiotensin-converting enzyme genotypes were identified using polymerase chain reaction. Blood pressure, plasma ACE activities, and plasma and erythrocyte zinc concentrations were measured at each prenatal clinical visit. Pregnancy outcome, the incidence of pregnancy-induced hypertension, and BP were not different among the three ACE genotypes. There was no significant correlation between plasma ACE activities and zinc concentrations. Zinc supplementation did not have a significant effect on either plasma ACE activities or zinc concentrations, probably because of the small sample size in our study. There was no effect of ACE gene polymorphism on pregnancy outcome, the incidence of pregnancy-induced hypertension, or changes in BP during pregnancy. Among each ACE genotype, plasma ACE activities did not correlate significantly with plasma zinc concentrations.


Class II major histocompatibility complex-restricted T cell function in CD4-deficient mice.
Previously, we and others have demonstrated that CD4-deficient mice have a normal number of T cells and B cells with a significant population of CD4-8-TcR alpha beta+ T cells. Surprisingly, however, these mice lacking CD4 show in vivo immunoglobulin isotype class switching from IgM to IgG in response to sheep erythrocytes and vesicular stomatitis virus. In this study we have depleted various subpopulations of T cells in vivo and shown that the population of CD4-8-TcR alpha beta+ T cells is responsible for providing "help" in the antibody response of CD4-deficient mice to vesicular stomatitis virus infection. We have used antigen-specific proliferation assays and blocking studies with class I and II major histocompatibility complex (MHC)-specific purified antibodies to show that these cells are class II MHC-restricted in responses against the T cell-dependent antigen keyhole limpet hemocyanin (KLH). Finally, phenotypic analysis of the CD4-CD8- thymocytes in CD4-deficient mice shows that these cells have a more mature phenotype than the CD4-8- thymocytes in wild type mice. These results indicate that CD4 is not absolutely necessary for positive selection or effector function of class II MHC-restricted helper T cells.


Theoretical modeling of charge trapping in crystalline and amorphous Al2O3.
The characteristics of intrinsic electron and hole trapping in crystalline and amorphous Al2O3 have been studied using density functional theory (DFT). Special attention was paid to enforcing the piece-wise linearity of the total energy with respect to electron number through the use of a range separated, hybrid functional PBE0-TC-LRC (Guidon et al 2009 J. Chem. Theory Comput. 5 3010) in order to accurately model the behaviour of localized states. The tuned functional is shown to reproduce the geometric and electronic structures of the perfect crystal as well as the spectroscopic characteristics of MgAl hole centre in corundum α-Al2O3. An ensemble of ten amorphous Al2O3 structures was generated using classical molecular dynamics and a melt and quench method and their structural characteristics compared with the experimental data. The electronic structure of amorphous systems was characterized using the inverse participation ratio method. Electrons and holes were then introduced into both crystalline and amorphous alumina structures and their properties calculated. Holes are shown to trap spontaneously in both crystalline and amorphous alumina. In the crystalline phase they localize on single O ion with the trapping energy of 0.38 eV. In amorphous phase, holes localize on two nearest neighbour oxygen sites with an average trapping energy of 1.26 eV, with hole trapping sites separated on average by about 8.0 Å. No electron trapping is observed in the material. Our results suggest that trapping of positive charge can be much more severe and stable in amorphous alumina rather than in crystalline samples.


Characterization of MSH2 variants by endogenous gene modification in mouse embryonic stem cells.
Mutations in the mismatch repair gene MSH2 underlie hereditary nonpolyposis colorectal cancer (Lynch syndrome). Whereas disruptive mutations are overtly pathogenic, the implications of missense mutations found in sporadic colorectal cancer patients or in suspected Lynch syndrome families are often unknown. Adequate genetic counseling of mutation carriers requires phenotypic characterization of the variant allele. We present a novel approach to functionally characterize MSH2 missense mutations. Our approach involves introduction of the mutation into the endogenous gene of murine embryonic stem cells (ESC) by oligonucleotide-directed gene modification, a technique we recently developed in our lab. Subsequently, the mismatch repair capacity of mutant ESC is determined using a set of validated functional assays. We have evaluated four clinically relevant MSH2 variants and found one to completely lack mismatch repair capacity while three behaved as wild-type MSH2 and can therefore be considered as polymorphisms. Our approach contributes to an adequate risk assessment of mismatch repair missense mutations. We have also shown that oligonucleotide-directed gene modification provides a straightforward approach to recreate allelic variants in the endogenous gene in murine ESC. This approach can be extended to other hereditary conditions.


Observing Proton Transfer Reactions Inside the MALDI Plume: Experimental and Theoretical Insight into MALDI Gas-Phase Reactions.
We evaluated the contribution of gas-phase in-plume proton transfer reactions to the formation of protonated and deprotonated molecules in the MALDI process. A split sample holder was used to separately deposit two different samples, which avoids any mixing during sample preparation. The two samples were brought very close to each other and desorbed/ionized by the same laser pulse. By using a combination of deuterated and non-deuterated matrices, it was possible to observe exclusively in-plume proton transfer processes. The hydrogen/deuterium exchange (HDX) kinetics were evaluated by varying the delayed extraction (DE) time, allowing the desorbed ions and neutrals to interact inside the plume for a variable period of time before being extracted and detected. Quantum mechanical calculations showed that the HDX energy barriers are relatively low for such reactions, corroborating the importance of gas-phase proton transfer in the MALDI plume. The experimental results, supported by theoretical simulations, confirm that the plume is a very reactive environment, where HDX reactions could be observed from 0 ns up to 400 ns after the laser pulse. These results could be used to evaluate the relevance of previously proposed (and partially conflicting) ionization models for MALDI. Graphical Abstract .


Donor variables affecting survival of autologous platelets.
While platelet survival studies have been carried out by numerous laboratories for many years, general agreement has not been reached on a single method or for the value of this test. Available data indicate that in spite of certain difficulties, reliable comparisons can be obtained using relatively small numbers of donors. Indeed, a difference in platelet survival of as little as 10 percent can be detected by using eight donors, whereas only four donors are required if one wishes to detect a 20 percent change. Should studies be done using donors preselected according to strict criteria of age, sex, health, etc.? It might be that less scatter among the data points would be observed and perhaps even fewer donors would be required to obtain significant values. In surveying the available information, it is clear that many investigators do have data on normal controls which could be extracted and analyzed to provide an answer to this question. Such information would be of great value in determining the criteria for selection of donors for autologous platelet survival studies.


Local adoptive transfer of skin delayed-type hypersensitivity initiated by a single T lymphocyte.
In vivo primed T cells injected in the footpad of naive recipients elicit a typical delayed-type hypersensitivity (DTH) reaction in the presence of their specific antigen. The values of the footpad swelling obtained after these transfers show a clear distinction between negative and positive responses. Serial dilutions of primed T cell populations allow the establishment of titration curves by limiting dilution analysis according to Poisson distribution. The single hit titration curve indicates that a unique cell type underlies the DTH reaction. Furthermore, by using cloned T cells it can be demonstrated that the transfer of a single cell is able to initiate a DTH reaction. The revelation of T cell activity at unit level needs an optimal dose of antigen mixed with the sampled cells.


Human herpesvirus 6 and cytomegalovirus in ileocolonic mucosa in inflammatory bowel disease.
Reactivation of a latent cytomegalovirus (CMV) may occur in inflammatory bowel disease (IBD). Data of human herpesvirus 6 (HHV-6)--a close relative to CMV--in active IBD are scarce. The aim of this study was to detect HHV-6 and CMV antigens in the mucosa of active and inactive IBD. 79 IBD patients (47 ulcerative colitis (UC) and 32 Crohn's disease (CD)) were recruited and endoscopic and histological disease activity was scored. Control group consisted of 15 non-IBD patients with normal colonoscopy. Immunohistochemical stainings for HHV-6B and CMV antigens were performed on biopsy specimens from the ileum and colorectum. The intensity of HHV-6B and CMV expression was graded as negative, mild, moderate, or intense. HHV-6B antigen was positive in 35 (44%) and CMV in 64 (81%). Of controls, 6 (40%) were mildly positive for HHV-6 and 6 (40%) for CMV. In IBD, both CMV and HHV-6B intensity correlated with endoscopic disease severity (CMV p = 0.010 and HHV-6 p = 0.048). Simultaneous HHV-6B and CMV antigen expression occurred in 29 (37%) and associated with endoscopic activity (p = 0.006) and to a number of immunosuppressives (p = 0.033). A significant difference in HHV-6B positivity was found between endoscopically active and inactive UC (p = 0.040). Both CMV and HHV-6B intensity correlated with histological severity in the rectal biopsy specimens (for CMV p = 0.040 and for HHV-6B p = 0.027). Both viruses occurred ubiquitously in the IBD mucosa. Coexistence of viruses was common and associated with disease activity and use of immunosuppressives. HHV-6B intensity correlated with endoscopic severity in UC.


Clinical symptoms in young adults with atypical chest pain attending the emergency department.
Clinical symptoms were studied in 69 consecutive patients below the age of 40 years who were attending the emergency unit because of unexplained chest pain. In a structured interview a few weeks after the emergency visit, only one-third of the patients reported that they believed in the doctor's diagnosis; they believed in a psychological or cardiac origin of the pain more often than the doctors. The chest pain was most often described as oppressive and/or stabbing. In 95% of cases it was central or left-sided. Associated symptoms were commonly reported, breathlessness being most commonly reported by two-thirds of the patients, followed by dizziness, palpitation and numbness/tingling. Mental symptoms such as tiredness, anxiety and tension were frequently reported. On the basis of the background literature the aetiology is discussed. We conclude that immediate symptom analysis, including psychosomatic symptoms, particularly breathing problems, is of central importance.


Serum thyrosinase in human lung carcinoma.
Compared to normal humans, lung carcinoma patients show increased tyrosinase activity. 7 serum enzymic fractions or carriers were present in the diseased state. Further, serum tyrosinase inhibitory factors generally were decreased in lung carcinoma patients compared to normal individuals.


Operable lung cancer associated with fever, anemia, and hepatomegaly.
A 26-year-old man, acutely ill with fever, anemia, and hepatomegaly, was found to have an epidermoid carcinoma by percutaneous needle biopsy of a left apical lung mass. Following resection of the necrotic tumor, fever, anemia, and hepatomegaly disappeared. The patient is alive with no evidence of cancer 30 months later.


Pharmacological modulation of HPA axis in depression - new avenues for potential therapeutic benefits.
One of the most consistent biological findings in major depression (MDD) is the altered activity of the hypothalamic-pituitary-adrenal (HPA) axis. It is not surprising that glucocorticoid receptor (GR), the common mechanism for stress-related changes in brain function, is a potential target of antidepressant drugs and therapies. All effective antidepressant treatments should trigger and maintain GR-related cellular processes necessary for recovery from MDD. Classic antidepressants act indirectly, by affecting the dynamic interplay between serotonin neurotransmission and HPA. On the other hand, certain compounds acting at supra-hypothalamic, HPA axis, glucocorticoid receptors, and post-receptor levels are being considered as new therapeutic options with the potential to modulate the aforementioned system in affective disorders directly. Different classes of drugs pharmacologically modify the HPA axis. This article summarizes the efficacy of classic antidepressants, as well as drugs classified as "antiglucocorticoids" (GR agonists, GR antagonists, dehydroepiandrosterone- DHEA, steroid synthesis inhibitors drugs, etc) in their capacity to heal glucocorticoid-mediated damage in depression. New avenues investigating the potential therapeutic benefits of antiglucocorticoids in affective disorders are at the proof-of-concept stage and future developments in this area deserve the full attention of psychiatrists and neuroscientists, as the current pharmacological treatment of MDD is far from perfect.


Mental health in deaf adults: symptoms of anxiety and depression among hearing and deaf individuals.
Individuals with disabilities encounter practical and social problems beyond those experienced by nondisabled individuals. This extra burden may in turn increase the risk of developing mental health problems. The objective of this article is to disclose the mental health situation among deaf individuals compared to a control sample of hearing individuals. The analyses are based on two separate Norwegian postal surveys, one among the general population (1995-1997) and one among the deaf population (2001). A shortened version of the Hopkins Symptom Checklist was used to disclose the degree of mental distress among the respondents. Three questions common to the studies were analyzed to determine differences between the two groups. Analyses revealed that the deaf respondents showed significantly more symptoms of mental health problems than the hearing respondents. The results point to the need for focussing more attention on the mental health of deaf children and adults. Society must be made aware of the special risks that deaf children and adults encounter with respect to mental health.


High-speed All-optical Modulator based on a Polymer Nanofiber Bragg grating printed by Femtosecond Laser.
On-chip optical modulator for high-speed information processing system has been widely investigated by majority of researchers but the connection with the fiber system is a difficulty. The fiber-based optical modulator is a good solution to this problem. Fiber Bragg Grating has good potential to be used as an optical modulator because of its linear temperature response, narrow bandwidth, and compact structure. In this paper, a new fiber-integrated all-optical modulator has been realized based on a polymer nanofiber Bragg grating printed by femtosecond laser. This device exhibits a fast temporal response of 176 ns and a good linear modulation of -45.43 pm/mW. Moreover, its stability has also been studied. This work firstly employs Bragg resonance to realize a fiber-integrated all-optical modulator and paves the way toward realization of multifunctional lab-in-fiber devices.


Interval of 9 h between birth of twins at term: case report and review of the literature.
We report the case of a primipara who delivered healthy twins vaginally at term with a time interval between twins of 9 h and 19 min. Neonatal outcome and further development were normal in both twins.


Combination of FASP and fully automated 2D-LC-MS/MS allows in-depth proteomic characterization of mouse zymogen granules.
Zymogen granule (ZG) constituents play important roles in pancreatic injury and disease. In previous studies, proteomic analyses with rat zymogen granules were separated by two-dimensional gel electrophoresis or one-dimensional SDS-PAGE, followed by in-gel tryptic digestion. In order to overcome the disadvantage of in-gel digestion and to carry out further in-depth proteomic analysis of the zymogen granules, in this study, by combining a filter-aided sample preparation method and fully automated 2D-LC-MS/MS technique, 800 ZG proteins were identified with at least two unique peptides for each protein, 75% of which have not been previously reported. The identified proteins revealed broad diversity in protein identity and function. This is the largest dataset of ZG proteome, and also the first dataset of the mouse ZG proteome, which may help elucidate on the molecular architecture of ZGs and their functions.


Intrathecal neostigmine, but not sympathectomy, relieves mechanical allodynia in a rat model of neuropathic pain.
Pain resulting from a usually nonpainful stimulus (allodynia) is a common characteristic of neuropathic pain. Among animal models of allodynia, tight ligature of lumbar spinal nerves has been of special interest because it has been reported to be relieved by sympathectomy. The purpose of this study was to determine whether spinal analgesic agents, which have opposite effects on sympathetic nervous system activity (clonidine decreases it and neostigmine increases it), have differing efficacy in this model. Male Sprague-Dawley rats were anesthetized, and the left L5 and L6 spinal nerves were ligated. At least 2 weeks later, a lumbar intrathecal or jugular intravenous catheter was inserted. Withdrawal threshold to mechanical stimulation of the hind paw was determined by application of von Frey filaments before surgery; after surgery; after intrathecal injection of clonidine, neostigmine, or their combination; after intravenous injection of phentolamine or guanethidine; and after surgical sympathectomy. Spinal nerve ligation reduced withdrawal threshold ipsilateral to the lesion. This allodynia was relieved by clonidine (50% block of allodynia at 20+/-1.2 microg and neostigmine (50% block of allodynia at 2+/-0.1 microg, and they interacted synergistically to block allodynia. Neither chemical nor surgical sympathectomy altered allodynia. These results disagree with previous observations that mechanical allodynia in this animal model depends on sympathetic nervous system activity. Therefore, intrathecally administered analgesic agents, one that reduces sympathetic outflow from the spinal cord (clonidine) and one that increases it (neostigmine), were similarly effective in this model.


Interactions of Ginkgo biloba extract (EGb 761), diazepam and ethyl beta-carboline-3-carboxylate on social behavior of the rat.
The social interaction test was used to examine the effects of an extract of Ginkgo biloba (EGb 761) and its possible interactions with diazepam and ethyl beta-carboline-3-carboxylate (beta-CCE). Pairs of naive (unfamiliar) male Wistar AF rats subjected to the same treatment were placed in a novel test arena that was brightly illuminated, and the duration (in s) of social contact was observed over a 10 min period. Single injections of EGb 761 (8-16 mg/kg, i.p.), given 30 min prior to testing, or repeated oral administration of the extract (48 or 96 mg/kg/day) for 8 days, significantly decreased social contact under conditions that did not influence locomotor activity. Injection of diazepam (1 mg/kg, i.p.), 30 min before testing, significantly increased social contact. Injection of diazepam to animals that had received repeated oral treatment with EGb 761 (96 mg/kg/ day) increased social interaction to an extent greater than observed with diazepam alone. Injection of beta-CCE (2-16 mg/kg, i.p.), 15 min before testing, significantly decreased social contact. When the animals were treated with EGb 761 (48 or 96 mg/kg/day, p.o. for 8 days) and beta-CCE (4 mg/kg), both of which decreased social interaction when administered alone, the resulting level of social contact was similar to that of control animals. Interactions with certain sites of central GABAA/ benzodiazepine/Cl- channel receptor complexes could be involved in mediating these effects of EGb 761, diazepam and beta-CCE.


Cost-effectiveness of hysteroscopy screening for infertile women.
This study assessed the cost-effectiveness of office hysteroscopy screening prior to IVF. Therefore, the cost-effectiveness of two distinct strategies - hysteroscopy after two failed IVF cycles (Failedhyst) and routine hysteroscopy prior to IVF (Routinehyst) - was compared with the reference strategy of no hysteroscopy (Nohyst). When present, intrauterine pathology was treated during hysteroscopy. Two models were constructed and evaluated in a decision analysis. In model I, all patients had an increase in pregnancy rate after screening hysteroscopy prior to IVF; in model II, only patients with intrauterine pathology would benefit. For each strategy, the total costs and live birth rates after a total of three IVF cycles were assessed. For model I (all patients benefit from hysteroscopy), Routinehyst was always cost-effective compared with Nohyst or Failedhyst. For the Routinehyst strategy, a monetary profit would be obtained in the case where hysteroscopy would increase the live birth rate after IVF by ≥ 2.8%. In model II (only patients with pathology benefit from hysteroscopy), Routinehyst also dominated Failedhyst. However, hysteroscopy performance resulted in considerable costs. In conclusion, the application of a routine hysteroscopy prior to IVF could be cost-effective. However, randomized trials confirming the effectiveness of hysteroscopy are needed.


[Doping in high-level athletes].
Recent doping affairs clearly demonstrate that all sports are concerned, leading to a generalized suspicion concerning champions and their performance. Many athletes struggling to reach their own personal goals have the unacceptable impression that they cannot compete fairly with competitors who are presumably (or effectively) doped. This situation may easily lead to deviant behavior. The sincerity of many high-level athletes when they discuss doping is very questionable in light of the notoriety and financial aspect linked with success. Public image is all important.Unfortunately, at this level is appears to be utopic to assume preventive measures would be effective. Several factors are involved Taking into consideration these different elements, one could be rather pessimistic concerning the eradication of doping in high-level athletes. There could even be a risk of seeing a trend towards public acceptance of more or less "tolerated" doping among professional athletes, similar to what is observed in the United States. It might be preferable to concentrate efforts on education and prevention in the young population.


Blocking of angiotensin II is more than blocking of transforming growth factor-beta.
Fibrosis is a common feature of chronic kidney diseases that is mediated by matrix-producing myofibroblasts. One potential origin of myofibroblasts is epithelial-mesenchymal transition (EMT) of tubuloepithelial cells. Transforming growth factor-beta (TGF-beta) is a key factor inducing EMT. Carvajal et al. demonstrate that angiotensin II induces EMT by classical stimulation of TGF-beta and also by a TGF-beta-independent pathway, both signaling via Smad molecules. Therefore, blockade of angiotensin II is more than lowering of blood pressure and inhibition of TGF-beta stimulation.


Analysis of the reversibility of biliary cirrhosis in young rats submitted to biliary obstruction.
Biliary atresia and other liver biliary obstructions are relevant conditions in pediatric surgery due to their progression to biliary cirrhosis and indication for liver transplantation. It is known that the period during which biliary obstruction persists determines the development of cirrhosis and its reversibility after a biliary drainage procedure. However, no time or histological markers of biliary cirrhosis reversibility have been established. One hundred and twenty-nine young Wistar rats underwent surgery for ligation of the common bile duct and were maintained until 8weeks. A part of these animals was submitted to biliary drainage surgery at 2, 3, 4, 5, or 6weeks after the initial procedure. After cyst formation at the site of obstruction, cyst-jejunal anastomosis was performed to restore bile flow. After biliary obstruction and drainage, liver samples were collected for histological and molecular analysis of the genes responsible for collagen deposition and fibrosis. The mortality rates were 39.8% and 56.7% after the first and second procedures, respectively. Ductular proliferation (p=0.001) and collagen deposition increased according to the period under obstruction (p=0.0001), and both alterations were partially reduced after biliary drainage. There were no significant differences in the values of desmin and α-actin according to the period during which the animal remained with biliary obstruction (p=0.09 and p=0.3, respectively), although increased values of transforming growth factor beta 1 (TGFβ1) occurred after 8weeks (p=0.000). Desmin levels decreased, and α-actin and TGFβ1 levels increased according to the period under obstruction. The molecular alterations were partially reversed after biliary drainage. The histologic and molecular changes in the liver parenchyma promoted by biliary obstruction in the young animal can be partially reversed by a biliary drainage procedure.


Comparative sensitivity of intracoronary injection of acetylcholine for the induction of coronary spasm in patients with various types of angina pectoris.
To elucidate the possible contribution of coronary artery spasm to the pathogenesis of angina pectoris, coronary arterial responses to intracoronary injection of acetylcholine were examined in patients with various types of angina pectoris. Coronary artery spasm with chest pain and/or electrocardiographic ischemic changes was angiographically demonstrated in 50 (85%) of 59 patients with angina pectoris. The sensitivity for coronary spasm was 92% (24 of 26) in patients with rest angina, 100% (16 of 16) in patients with both rest and effort angina, and 59% (10 of 17) in patients with effort angina, while it was only 6% (1 of 16) in patients without coronary artery disease. When patients with effort angina were subdivided according to the variability of anginal threshold for exertional angina, the sensitivity for coronary spasm was as high as 90% (9 out of 10) in patients with variable-threshold angina. In contrast, coronary spasm was less frequently (p less than 0.05) induced in patients with fixed-threshold angina (1 of 7, 14%). These results suggest that coronary arteries in patients with angina pectoris are quite susceptible to acetylcholine except in those patients with stable exercise tolerance or anginal threshold. Thus coronary artery spasm appears to play a significant role for the pathogenesis of angina pectoris in a large proportion of patients with effort angina as well as in patients with rest angina.


[Stability analysis of on-board calibration system of spatially modulated imaging Fourier transform spectrometer].
Spatially modulated imaging Fourier transform spectrometer (SMIFTS) was an instrument that depended on interference, and after calibration, the reconstruction spectrum can quantificationally reflect the diffuse reflection of target under sunshine. On-board calibration of SMIFTS confirmed the change of SMIFTS according to relative spectrum calibration, inspected long-time attenuation of SMIFTS optical system, and corrected export data of SMIFTS. According to the requirement of remote sensor application, it must stay in vacuum environment for a long time. As a radiant standard, the stability of lamp-house in long time is the most important characteristic of on-board calibration system. By calculation and experimentation, analyses of on-board calibration of SMIFTS, and testing spaceflight environment characteristic of on-board calibration, the difficulty in the key parts of on-board calibration of SMIFTS such as lamp-house, spectrum filter and integrating sphere was solved. According to the radiation-time stability testing for lamp-house and optics, particle-radiation testing, environmental-mechanics testing and hot vacuum examination, good result was obtained. By whole-aperture and part-field comparative wavelength calibration, the spectrum curve and before-launch interferogram were obtained. After comparison with reconstruction spectrum under different condition, the stability and credibility of SMIFTS on-board calibration system was proved.


1-(Benzenesulfonyl)-1,5-dihydro-4,1-benzoxazepine as a new scaffold for the design of antitumor compounds.
Bozepinib is a potent and selective anticancer compound which chemical structure is made up of a benzofused seven-membered ring and a purine moiety. We previously demonstrated that the purine fragment does not exert antiproliferative effect per se. A series of 1-(benzenesulfonyl)-4,1-benzoxazepine derivatives were synthesized in order to study the influence of the benzofused seven-membered ring in the biological activity of bozepinib by means of antiproliferative, cell cycle and apoptosis studies. Our results show that the methyleneoxy enamine sulfonyl function is essential in the antitumor activity of the structures and thus, it is a scaffold suitable for further modification with a view to obtain more potent antitumor compounds.


Polyvinylidene fluoride/silane-treated hydroxyapatite mixed matrix membrane for enzyme capturing.
The silane coupling agent, N-(β-aminoethyl)-γ-aminopropyltrimethoxy silane (KH792), was employed to modify the surfaces of nano-hydroxyapatite (HAP) particles. The mixed matrix membranes (MMMs) were prepared by embedding pure HAP and HAP modified with KH792 (KH792-HAP) inside polyvinylidene fluoride (PVDF) matrix respectively. The MMMs were further characterized concerning permeability and adsorption capacity. Langmuir adsorption isotherm provides better fit for HAP and KH792-HAP than Freundlich isotherm. KH792-HAP has better distribution in the polymeric matrix compared to HAP in the polymeric matrix. The MMMs showed purification of enzyme via static adsorption and dynamic adsorption, and showed the potential of using MMMs for enzyme capturing in enzyme purification techniques. The lysozyme (LZ) was used as a model enzyme. The properties and structures of MMMs prepared by immersion phase separation process were characterized by pure water flux, LZ adsorption and scanning electron microscopy (SEM).


Nongastric H-K-ATPase in rodent prostate: lobe-specific expression and apical localization.
The molecular basis of active ion transport in secretory glands such as the prostate is not well characterized. Rat nongastric H-K-ATPase is expressed at high levels in distal colon surface cell apical membranes and thus is referred to as "colonic." Here we show that the ATPase is expressed in rodent prostate complex in a lobe-specific manner. RT-PCR and Western blot analyses indicate that rat nongastric H-K-ATPase alpha-subunit (alpha(ng)) mRNA and protein are present in coagulating gland (anterior prostate) and lateral and dorsal prostate and absent from ventral lobe, whereas Na-K-ATPase alpha-subunit is present in all lobes. RT-PCR analysis shows that Na-K-ATPase alpha(4) and alpha(3) and gastric H-K-ATPase alpha-subunit are not present in significant amounts in all prostate lobes. Relatively low levels of Na-K-ATPase alpha(2) were found in lateral, dorsal, and anterior lobes. alpha(ng) protein expression is anteriodorsolateral: highest in coagulating gland, somewhat lower in dorsal lobe, and even lower in lateral lobe. Na-K-ATPase protein abundance has the reverse order: expression in ventral lobe is higher than in coagulating gland. alpha(ng) protein abundance is higher in coagulating gland than distal colon membranes. Immunohistochemistry shows that in rat and mouse coagulating gland epithelium alpha(ng) protein has an apical polarization and Na-K-ATPase alpha(1) is localized in basolateral membranes. The presence of nongastric H-K-ATPase in rodent prostate apical membranes may indicate its involvement in potassium concentration regulation in secretions of these glands.


Skin collagen reproduction increased by ascorbic acid derivative iontophoresis by frequent-reversal bipolar electric stimulation.
The effect of the iontophoresis of ascorbic acid (Vitamin C; VC) derivative with frequent-reversal bipolar electric stimulation on the production of collagen in rat skin was evaluated in terms of hydroxyproline content through high-performance liquid chromatography. First, a control group was not given electrical stimulation and four groups were stimulated with a unipolar pulse for 0.5-10 min every day for one week. The hydroxyproline level in the skin was increased depending on the length of the stimulation. Second, a control group was not given any electrical stimulation, and three groups were treated with (a) VC solution without any stimulation, (b) a bipolar pulse for 10 min with saline, or (c) a bipolar pulse for 5 min with the VC solution. Significant increases were found in all the stimulation groups, although these treated with the VC solution without any stimulation did not have any effects compared to the control. Thus, in order to increase the hydroxyproline levels in skin, a VC must be delivered with bipolar stimulation as a method of iontophoresis. These results suggest that our newly developed electric stimulation is effective at increasing skin collagen content, and that bipolar stimulation is more effective on the iontophoresis of not only VC but also some medicines such as low- and high-molecular drugs directed to the target organ (7).


Corneal topographic analysis after excimer photorefractive keratectomy.
To evaluate the corneal surface changes and visual quality after excimer photorefractive keratectomy (PRK) for myopia. Corneal topographic analysis was performed on 23 patients (38 eyes) with myopia after PRK at the 1-, 3-, 6- month postoperative follow-up visits. The corneal sphericity was changed after excimer PRK. As time went on, the mean surface regularity index (SRI) and the surface asymmetry index (SAI) decreased gradually, and the corneal surface became more smooth; mean simulated keratoscope readings (SimK) showed a gradual restoration. At the 6-month postoperative examination, corneal topography showed four main patterns of ablation: round or oblong (50%), collar-button (23.68%), semicircular (18.42%) and central island (7.9%). Patterns of ablation were correlated with visual acuity. The quantitative analysis of corneal topography is essential for evaluating corneal surface changes after PRK and helpful in the surgical design of PRK and in predicting the refractive outcome with greater precision.


Chinese pediatric Tuina on children with acute diarrhea: study protocol for a randomized sham-controlled trial.
Acute pediatric diarrhea is one of the most common causes of morbidity and mortality worldwide and seriously affects the health of children. Previous studies have shown that pediatric Tuina, a traditional Chinese medicine therapy, has potential therapeutic benefits for acute pediatric diarrhea. However, the evidence for its effectiveness is insufficient due to the lack of high-quality clinical studies. Our aim is to evaluate the efficacy of Chinese pediatric Tuina for children aged 0-6 years with acute diarrhea. This study is a randomized, double-blind, sham-controlled trial. We will include 122 children with acute diarrhea from Dongguan Kanghua Hospital in Guangdong province, China. The patients will be allocated into either the pediatric Tuina group or the sham Tuina group in a 1:1 ratio. The treatment will last for 3 days followed by an 11-day follow-up period. Both groups will receive usual care. In addition, the experimental group will receive 15-25 min of Chinese pediatric Tuina, while the control group will receive 15-25 min of sham pediatric Tuina. Both groups will receive treatments once per day, for 3 consecutive days. Primary outcome measures are diarrhea days from baseline and diarrhea times on the third day. Secondary outcome measures are the global change rating and period of days when the stool character changes to normal. Safety assessments will be monitored during each visit. This clinical trial is designed to evaluate the efficacy of pediatric Tuina for children with acute diarrhea. We expect results to provide solid evidence and support for pediatric Tuina as an appropriate treatment for children with acute diarrhea. ClinicalTrials.gov, NCT03005821. Registered on 29 December 2016.


Beyond The Therapeutic Effect of rTMS in Alzheimer's Disease: A Possible Neuroprotective Role of Hippocampal BDNF? : A Minireview.
Repetitive transcranial magnetic stimulation (rTMS) is a well known non-invasive brain stimulation procedure which is capable of inducing the expression of the hippocampal BDNF that has been already shown to exert significant neuroprotective and pro-cognitive effects in AD. However, it is nearly impossible directly to evaluate the BDNF expression in humans after rTMS application. Here we summarized the underlying mechanisms of the neuroprotective and procognitive effect of BDNF that can be induced through a region-specific rTMS approach. Additionally, we have also evaluated the role of Magnetic Resonance Spectroscopy in monitoring the BDNF response after rTMS application in Alzheimer's Disease. We have provided strong evidence that rTMS exerts significant neuroprotective and pro-cognitive effects through the expression of hippocampal BDNF. Furthermore, Magnetic Resonance Spectroscopy might play a critical role in monitoring the BDNF response after rTMS application in AD patients. Such a sophisticated approach might be able to enlighten us on the time-dependent cognitive and neuroprotective correlates of the rtMS application in AD patients.


Two-phase anaerobic digestion of lignocellulosic hydrolysate: Focusing on the acidification with different inoculum to substrate ratios and inoculum sources.
Biogas production from lignocellulosic hydrolysate is of great potential for lignocellulosic materials. Two-phase anaerobic digestion was proposed in this study. Acidogenic fermentation was carried out with corn straw hydrolysate as feedstock for volatile fatty acids (VFAs) production. Using anaerobic sludge (AnS), different inoculum to substrate ratios (ISRs) of 0.5:1, 1:1 and 2:1 were investigated. The highest VFAs yield was obtained at ISR of 0.5:1.VFAs composition analysis showed that butyric acid was the predominant acid, followed by acetic acid and propionic acid. The effects of AnS and aerobic sludge (AeS) on the acidogenic performance of hydrolysate were compared. The optimum VFA yields were 0.38 g/g COD-added for AnS and 0.32 g/g COD-added for AeS with HRT of 5 d, respectively. The bacterial diversities of inocula and digestates were analyzed by high-throughput sequencing. Two origins of inocula had distinct bacterial structures, but they did share core communities that included Firmicutes, Chloroflexi, Proteobacteria and Bacteroidetes at phylum level. The bacterial communities of both digestates changed significantly as compared with those in inoculum. Firmicutes was absolutely dominant in all the bacterial species. Therefore, the AeS could be an option as the acidogenic inoculum. The microbial information will be beneficial for the enrichment and acclimatization of microbes. In methanogenic process, VFAs obtained in acidogenic stage could be efficiently converted into methane. The ultimate methane yield at organic loading rate (OLR) of 8 g/L·d could reach 290 mL CH4/g COD-added and 279 mL CH4/g COD-added for AnS and AeS acidified digestate. Two-phase anaerobic digestion was proved to be suitable for bioconversion of lignocellulosic hydrolysate into biogas.