Electrophysiological effects of phencyclidine in the medial prefrontal cortex of the rat.
The effects of local applications of phencyclidine (PCP) and dopamine (DA) on neurons of the medial prefrontal cortex were investigated using single unit recording techniques. The activity of the majority of cells in the deeper layers of the medial prefrontal cortex was depressed by both phencyclidine and DA, whereas increases, as well as decreases, in the firing rates were observed in cells located in the superficial cortical layers. The stereospecificity of the responses of deeper cells to phencyclidine was demonstrated using the enantiomers of 1-(-1-phenylcyclohexyl)-3-methylpiperidine (PCMP). Phencyclidine was found to be 1.5 times more potent than (+) PCMP and 3 times more potent than (-) PCMP. Finally, the DA receptor antagonist fluphenazine, blocked the phencyclidine-elicited depressions of unit activity in the deep prefrontal cortex. Taken together, the data indicate that the DA-like effects of phencyclidine on neurons of the medial prefrontal cortex are mediated by DA receptors and provide pharmacological support for the idea that psychomotor stimulant drugs have specific actions on targets of the ventral tegmental area (A10) dopamine system.


Concepts of burden in giving care to older relatives: a study of female caregivers in a Mexico City neighborhood.
This study examined how women in a Mexico City suburb conceptualized the construct of burden within the context of giving care to older relatives. Data were collected on 41 women through semi-structured interviews regarding their caregiving experiences to elderly relatives. Phenomenological principles were used to analyze meanings and understandings of caregiving experiences. Burden was a multi-dimensional construct that referred to specific situations that made caregivers feel emotionally or physical "heavy." Burden also referred to "being a burden" by being in the way, making things difficult, or being a 'weight' on caregivers' shoulders. However, women in this study also viewed burden as a positive sacrifice that involved love, initiative, and good will. This study is an important first step in defining the ways in which caregiving is positively and negatively meaningful for Mexican caregivers and their families. Our findings offer an additional dimension of caregiver burden to broaden our understanding and measurement of the construct. The development of culturally appropriate instruments to measure caregiving burden in Mexico is necessary to provide an empirical foundation for policy recommendations that address the growing need for institutional support of caregivers. Moreover, our findings suggest that researchers studying caregiving in Mexico should think about how burden is measured on existing instruments before adapting them for widespread use.


Microscopic and ultrastructural evidences in human skin following calcium hydroxylapatite filler treatment.
This study uses light and electron microscopes to gain a better knowledge of the interactions of calcium hydroxylapatite filler with the connective tissue of the skin and the modifications of the human deep dermis, after 2 months of treatment. Some morphological evidences of this observational study of filler treated tissue support-specific mechanism involved in the structural modifications of both filler microspherules and cells of the connective tissue. They demonstrate the absence of any immunological reaction and show that the used filler is modified very slowly over time by the action of cells of the connective tissue closely related to the filler without any activity of phagocytosis. Furthermore, associated with the modifications of the filler, evidences of stimulatory effects on dermal fibroblasts are reported.


The relationship between Helicobacter pylori infection, the virulence genotypes of the infecting strain and gastric cancer in the African setting.
The relationship between Helicobacter pylori infection and gastric carcinoma remains controversial, especially in the African setting where infection is common, while gastric cancer is perceived to be uncommon, the basis of the so called 'African enigma'. This discrepancy between infection and the development of disease is commonly attributed to differences in host, environment and bacterial factors. Interest in the bacterial factors has focused on heterogeneity in the so-called 'virulence genes'. The aim of this prospective, case-controlled study was to establish whether H. pylori infection is significantly associated with gastric cancer and to investigate whether gastric cancer is associated with genotypically distinct (as it relates to the candidate virulence genes) organisms in this population. Patients with histologically confirmed gastric cancer were matched with nonulcer dyspeptic controls for age (within 5 years), gender and ethnicity. Helicobacter pylori status was determined by RUT, histology, culture and serology (locally validated and used as default determinant of H. pylori status). Tumors were classified according to the Lauren classification. The 'virulence genotype' of 17 paired culture samples was determined by previously described and validated molecular techniques (cagA presence, vacA alleles, structure of the cag pathogenicity island and analysis of the iceA alleles). Categorical variables were analysed by the chi2 test. Forty-eight patients (median age 59 years) could be adequately matched to controls. 39/48 (81%) cases and 43/48 (90%) controls were H. pylori positive (NS). Significant differences in the virulence genotypes of infecting strains were noted: vacAs2-controls 24%, cases 0%, p < .00001; vacAs1 present - cases 100%, controls 76%, p < .05; cagA-3'-length > 650 bp - cases 47%, controls 0%, p < .002; cag pathogenicity island intact - cases 82%, controls 43%, p < .04; iceA1 - cases 53%, controls 6%, p < .005. cagA was found in all subjects. This study indicates that, in this African population at least, there is no difference in the prevalence of H. pylori infection when comparing gastric cancer cases with matched controls. However, the findings suggest that gastric cancer may be associated with infection by organisms that are genotypically different from those not associated with disease.


Cerebrospinal fluid alterations of the serotonin product, 5-hydroxyindolacetic acid, in neurological disorders.
Although patients with low cerebrospinal fluid (CSF) serotonin metabolite levels have been reported, inborn errors of the rate-limiting enzyme of serotonin synthesis (tryptophan hydroxylase, TPH) have not been described so far. In this study we aimed to evaluate CSF alterations of the serotonin metabolite 5-hydroxyindolacetic acid (5-HIAA) in patients with neurological disorders and to explore a possible TPH deficiency in some of them. A total of 606 patients (286 males, 320 females, mean age 4 years and 6 months, SD 5 years and 7 months) underwent CSF analysis of neurotransmitter metabolites by reverse phase high performance liquid chromatography. Results were compared with values established in a control population. Patients' medical records were reviewed to determine diagnosis and clinical features. A primary defect of biogenic amines was genetically investigated in indicated patients. Low 5-HIAA was seen in 19.3%. Of these, 22.2% showed inborn errors of metabolism (mitochondrial disorders being the most frequent at 10.2% of low 5-HIAA patients) and neurogenetic conditions. Other relatively frequent conditions were pontocerebellar hypoplasia (4.3%), Rett syndrome (4.3%), and among congenital nonetiologically determined conditions, epilepsy including epileptic encephalopathies (26.4%), leukodystrophies (6.8%), and neuropsychiatric disturbances (4.2%). Mutational analysis of the TPH2 gene, performed in five candidate patients, was negative. Although frequency of secondary alteration of 5-HIAA was relatively high in patients with neurological disorders, this finding was more frequently associated with some neurometabolic disorders, epileptic encephalopathies, and neuropsychiatric disturbances. No inborn errors of TPH were found. Due to serotonin's neurotrophic role and to ameliorate symptoms, a supplementary treatment with 5-hydroxytriptophan would seem advisable in these patients.


Targeted and untargeted quantification of quorum sensing signalling molecules in bacterial cultures and biological samples via HPLC-TQ MS techniques.
Quorum sensing (QS) is the ability of some bacteria to detect and to respond to population density through signalling molecules. QS molecules are involved in motility and cell aggregation mechanisms in diseases such as sepsis. Few biomarkers are currently available to diagnose sepsis, especially in high-risk conditions. The aim of this study was the development of new analytical methods based on liquid chromatography-mass spectrometry for the detection and quantification of QS signalling molecules, including N-acyl homoserine lactones (AHL) and hydroxyquinolones (HQ), in biofluids. Biological samples used in the study were Pseudomonas aeruginosa bacterial cultures and plasma from patients with sepsis. We developed two MS analytical methods, based on neutral loss (NL) and product ion (PI) experiments, to identify and characterize unknown AHL and HQ molecules. We then established a multiple-reaction-monitoring (MRM) method to quantify specific QS compounds. We validated the HPLC-MS-based approaches (MRM-NL-PI), and data were in accord with the validation guidelines. With the NL and PI MS-based methods, we identified and characterized 3 and 13 unknown AHL and HQ compounds, respectively, in biological samples. One of the newly found AHL molecules was C12-AHL, first quantified in Pseudomonas aeruginosa bacterial cultures. The MRM quantitation of analytes in plasma from patients with sepsis confirmed the analytical ability of MRM for the quantification of virulence factors during sepsis. Graphical abstract.


Behavior of Steel-Sheathed Shear Walls Subjected to Seismic and Fire Loads.
A series of tests was conducted on six 2.7 m × 3.7 m shear wall specimens consisting of cold-formed steel framing sheathed on one side with sheet steel adhered to gypsum board and on the opposite side with plain gypsum board. The specimens were subjected to various sequences of simulated seismic shear deformation and fire exposure to study the influence of multi-hazard interactions on the lateral load resistance of the walls. The test program was designed to complement a parallel effort at the University of California, San Diego to investigate a six-story building subjected to earthquakes and fires. The test results reported here indicate that the fire exposure caused a shift in the failure mode of the walls from local buckling of the sheet steel in cases without fire exposure, to global buckling of the sheet steel with an accompanying 35 % reduction in lateral load capacity after the wall had been exposed to fire. This behavior appears to be predictable, which is encouraging from the standpoint of residual lateral load capacity under these severe multi-hazard actions.


Fatal haemorrhage due to extensive fragility of medium- and large-sized arteries and veins in a young patient with neurofibromatosis 1.
Neurofibromatosis type 1 (NF1) is one of the most frequently inherited diseases. Vascular manifestations have been reported, mostly concerning stenosis of the renal artery or spontaneous rupture of single arteries. Also, venous aneurysms have been reported. We present clinical and pathological findings of a young patient with NF1 with lethal bleeding due to spontaneous rupture of a lumbal and renal artery. High fragility of several arteries as well as veins in the abdominal and pelvic arterial and venous system resulted in a destroyed vessel structure caused by invading neurogen fibres. In rare cases, NF1 is associated with a severe systemic vasculopathy concerning the arterial and venous vascular system.


Can a Therapeutic Strategy for Hypotension Improve Cerebral Perfusion and Oxygenation in an Experimental Model of Hemorrhagic Shock and Severe Traumatic Brain Injury?
Restoration of brain tissue perfusion is a determining factor in the neurological evolution of patients with traumatic brain injury (TBI) and hemorrhagic shock (HS). In a porcine model of HS without neurological damage, it was observed that the use of fluids or vasoactive drugs was effective in restoring brain perfusion; however, only terlipressin promoted restoration of cerebral oxygenation and lower expression of edema and apoptosis markers. It is unclear whether the use of vasopressor drugs is effective and beneficial during situations of TBI. The objective of this study is to compare the effects of resuscitation with saline solution and terlipressin on cerebral perfusion and oxygenation in a model of TBI and HS. Thirty-two pigs weighing 20-30 kg were randomly allocated into four groups: control (no treatment), saline (60 ml/kg of 0.9% NaCl), terlipressin (2 mg of terlipressin), and saline plus terlipressin (20 ml/kg of 0.9% NaCl + 2 mg of terlipressin). Brain injury was induced by lateral fluid percussion, and HS was induced through pressure-controlled bleeding, aiming at a mean arterial pressure (MAP) of 40 mmHg. After 30 min of circulatory shock, resuscitation strategies were initiated according to the group. The systemic and cerebral hemodynamic and oxygenation parameters, lactate levels, and hemoglobin levels were evaluated. The data were subjected to analysis of variance for repeated measures. The significance level established for statistical analysis was p < 0.05. The terlipressin and saline plus terlipressin groups showed an increase in MAP that lasted until the end of the experiment (p < 0.05). There was a notable increase in intracranial pressure in all groups after starting treatment for shock. Cerebral perfusion pressure and cerebral oximetry showed no improvement after hemodynamic recovery in any group. The groups that received saline at resuscitation had the lowest hemoglobin concentrations after treatment. The treatment of hypotension in HS with saline and/or terlipressin cannot restore cerebral perfusion or oxygenation in experimental models of HS and severe TBI. Elevated MAP raises intracranial pressure owing to brain autoregulation dysfunction caused by TBI.


The physical and genetic organization of a viral genome.
Mutants of SV40 with deletions of a few to several thousand base pairs have been constructed in vitro and cloned in cultured monkey cells. The location and size of these deletions has been determined by restriction endonuclease mapping and electron microscopic and enzymatic analysis of DNA heteroduplex molecules. Analysis of the phenotype of these deletion mutants permits us to specify the locations of the known SV40 genes, in particular, the novel organization of SV40s two early genes that are required for oncogenesis.


Lung Carcinogenesis by Diesel Exhaust Particles and the Carcinogenic Mechanism Via Active Oxygens.
In an experiment to clarify the involvement of oxygen radicals in lung carcinogenesis induced by diesel exhaust particles (DEP), we found that there is a strong relation between lung tumor response and formation of 8-hydroxydeoxyguanosine (8-OHdG) in lung DNA of mice administered DEP by repeated intratracheal instillation. Repeated intratracheal instillation of DEP also induced the activity of cytochrome P-450 reductase in the lungs as a representative enzyme of superoxide generation, and two types of nitric oxide (NO) synthase, cNOS and iNOS, in the lungs. On the other hand, activities of CuZn-superoxide dismutase (SOD) and Mn-SOD antioxidant enzymes were depressed by the instillation of DEP. These results suggest that generation of superoxide, hydroxyI radical, and nitric oxide are increased in epithelial cells in airways, and that the increased superoxide and nitric oxide react very easily to produce peroxynitrite (ONOO(-)). The peroxynitrite also produce hydroxyI radical. The hydroxyl radical may play an important role in carcinogenesis by DEP.


Preparation and characterization of slow-release fertilizer encapsulated by biochar-based waterborne copolymers.
To enhance the effectiveness of polyvinyl alcohol (PVA) coated fertilizers, a novel slow-release fertilizer was developed using biochar and waterborne copolymer of PVA and polyvinylpyrrolidone (PVP) as coating materials. The effects of botanical origins and doses of biochar as well as copolymer concentrations on biochar-copolymer structures and properties were investigated. Our results indicated that the water absorbency of blend films differing in biochar origins showed different responses to these three factors. Generally, biochar decreased water absorbency of copolymer with an increased degradability, and contributed to improving the slow-release property of coated urea. In particular, rice biochar-based copolymer (S5 film) had less hydrophilic OH bonds and encapsulated urea granules more compactly and densely. The urea particles coated with rice biochar-based copolymer (S5 film) exhibited an excellent release behavior of 65.28% nutrient leaching on the 22th day. Therefore, this work has demonstrated the potential of biochar-based copolymer from different botanical biochar for improving the effectiveness of fertilizers.


Procedure for characterization of creatine kinase variants on agarose electrophoretograms.
Electrophoretograms of atypical creatine kinase (EC 22.214.171.124, CK) isoenzymes may be difficult to interpret. In addition, their presence may cause discrepancies between results by immunological determinations for mass or activity and those by electrophoresis. Here we outline procedures to use in characterizing apparently atypical CK isoenzymes observed after electrophoresis, as illustrated by a CK isoenzyme variant that migrated as CK-MB.


Factors associated with psychosocial adjustment in newly diagnosed young to middle-aged women with breast cancer: A cross-sectional study.
To examine the psychosocial adjustment of young to middle-aged women who were newly diagnosed with breast cancer and to determine the comprehensive risk factors contributing to psychosocial adjustment. This study was carried out on 358 young to middle-aged women who recently received a breast cancer diagnosis in two hospitals in Guangzhou, China. Participants reported data about sociodemographic characteristics, disease and treatment information, coping modes, social support, self-efficacy, and psychosocial adjustment. To analyze the data, the researchers utilized independent t-tests, one-way analysis of variance, and multiple linear regression. The results showed that the participants exhibited a moderate level of psychosocial maladjustment, with a mean score of 42.44 ± 15.38. Additionally, 30.4% of the participants were classified as having severe psychosocial maladjustment. The study identified the coping mode of acceptance-resignation (β = 0.367, P < 0.001), mode of avoidance (β = -0.248, P = 0.001), social support (β = -0.239, P < 0.001), self-efficacy (β = -0.199, P = 0.001) as factors that impacted the level of psychosocial adjustment. Psychosocial adjustment among young to middle-aged women who were newly diagnosed with breast cancer is affected by self-efficacy, social support, and coping modes. Healthcare professionals should pay attention to psychosocial adjustment in young to middle-aged women with breast cancer at the time of diagnosis, and could formulate effective interventions to improve their psychosocial adjustment by increasing self-efficacy, promoting social support, and encouraging effective coping.


Dual-light control of nanomachines that integrate motor and modulator subunits.
A current challenge in the field of artificial molecular machines is the synthesis and implementation of systems that can produce useful work when fuelled with a constant source of external energy. The first experimental achievements of this kind consisted of machines with continuous unidirectional rotations and translations that make use of 'Brownian ratchets' to bias random motions. An intrinsic limitation of such designs is that an inversion of directionality requires heavy chemical modifications in the structure of the actuating motor part. Here we show that by connecting subunits made of both unidirectional light-driven rotary motors and modulators, which respectively braid and unbraid polymer chains in crosslinked networks, it becomes possible to reverse their integrated motion at all scales. The photostationary state of the system can be tuned by modulation of frequencies using two irradiation wavelengths. Under this out-of-equilibrium condition, the global work output (measured as the contraction or expansion of the material) is controlled by the net flux of clockwise and anticlockwise rotations between the motors and the modulators.


The effects of oxygen radicals on the activity of nitric oxide synthase and guanylate cyclase.
Reactive oxygen species such as superoxides, hydrogen peroxide (H2O2) and hydroxyl radicals have been suggested to be involved in the catalytic action of nitric oxide synthase (NOS) to produce NO from L-arginine. An examination was conducted on the effects of oxygen radical scavengers and oxygen radical-generating systems on the activity of neuronal NOS and guanylate cyclase (GC) in rat brains and NOS from the activated murine macrophage cell line J774. Catalase and superoxide dismutase (SOD) showed no significant effects on NOS or GC activity. Nitroblue tetrazolium (NBT, known as a superoxide radical scavenger) and peroxidase (POD) inhibited NOS, but their inhibitory actions were removed by increasing the concentration of arginine or NADPH respectively, in the reaction mixture. NOS and NO-dependent GC were inactivated by ascorbate/FeSO4 (a metal-catalyzed oxidation system), 2'2'-azobis-amidinopropane (a peroxy radical producer), and xanthine/xanthine oxidase (a superoxide generating system). The effects of oxygen radicals or antioxidants on the two isoforms of NOS were almost similar. However, H2O2 activated GC in a dose-dependent manner from 100 microM to 1 mM without significant effects on NOS. H2O2-induced GC activation was blocked by catalase. These results suggested that oxygen radicals inhibited NOS and GC, but H2O2 could activate GC directly.


Mental health of pregnant women during the COVID-19 pandemic: A longitudinal study.
Several studies have reported the susceptibility of pregnant women to emotional instability and stress. Thus, pregnancy may be a risk factor that could deepen the already negative effects of the current COVID-19 pandemic. Therefore, the aim of this study is to analyze longitudinally the psychopathological consequences of the pandemic in pregnant women, and to explore differences with non-pregnant women. The participants in this study were 102 pregnant women, and a control group of 102 non-pregnant women (most of them reported having university studies and little financial impact from the pandemic). They completed the Beck Depression Inventory-II, the State-Trait Anxiety Inventory, and the Positive and Negative Affect Schedule, in three different times (2, 14, and 47 days after the start of the lockdown). In a time range of 50 days of quarantine, all women showed a gradual increase in psychopathological indicators and a decrease in positive affect. Pregnant women showed a more pronounced increase in depression, anxiety and negative affect than the non-pregnant women did. In addition, pregnant women showed a more pronounced decrease in positive affect. It is important for institutions dedicated to perinatal health care to count on empirical information to optimize the provision of their services.


Gonadal steroids and neuropeptide Y-opioid-LHRH axis: interactions and diversities.
We report that the two classes of regulatory neuropeptides, neuropeptide Y (NPY) and endogenous opioid peptides (EOP), modulate luteinizing hormone (LH) release in diverse fashion in gonad-intact rats. Each neuropeptide acts at two loci, the hypothalamus and pituitary, to excite (NPY) or inhibit (EOP) LH release. At the hypothalamic level, NPY stimulates luteinizing hormone releasing hormone (LHRH) release, a response mediated by alpha 2-adrenoreceptors and amplified in the presence of adrenergic agonists. At the pituitary level, NPY acts in concert with LHRH to amplify the LH response. In contrast, EOP inhibit LHRH release by decreasing the supply of excitatory adrenergic signals in the vicinity of LHRH neurons in the preoptic-tuberal pathway, and at the pituitary level, they decrease LH release in response to LHRH. Further, the gonadal steroidal milieu facilitates NPY neurosecretion and postsynaptic expression of NPY in concert with adrenergic system; a similar clear-cut facilitatory effect of gonadal steroids on EOP secretion is not yet obvious. Our additional studies imply that the EOP system has the potential to increase sensitivity towards gonadal steroids and that to induce the preovulatory LH surge the neural clock may decrease the inhibitory EOP tone prior to the critical period on proestrus. This antecedent neural event allows the excitatory adrenergic and NPY signals to evoke LHRH secretion at a higher frequency approximating that seen in ovariectomized rats. Further studies are under way to delineate the steroid-induced subcellular events that integrate the action of these regulatory peptides in the control of the episodic LHRH secretion pattern which sustains basal and cyclic gonadotropin release in the rat.


Cloning and nucleotide sequence of senN, a novel 'Bacillus natto' (B. subtilis) gene that regulates expression of extracellular protein genes.
A new 'Bacillus natto' gene, senN, that regulates the expression of several extracellular proteins in B. subtilis has been cloned and sequenced. senN codes for a small, highly basic protein with an amino acid sequence different from the products coded by the regulatory genes sacQ, sacV, prtR and hpr. SenN stimulates gene expression at the transcriptional level. A putative homologous locus has been detected in the B. subtilis chromosome by Southern blotting.


Responses of human sling and clasp fibers to cholecystokinin (CCK) and gastrin through CCK receptors.
Cholecystokinin (CCK) and gastrin exert their influences via CCK receptors. This research was conducted to look at the responses of the sling and clasp fibers forming the human lower esophageal sphincter (LES) to CCK and gastrin, and the role of CCK receptors in the responses. Muscle strips of sling and clasp fibers from the LES were obtained from patients undergoing subtotal esophagectomy. Isometric tension responses of the strips to CCK-8 and gastrin-17 were studied, and the maximum effect (E(max)) for each agonist was derived. CCK-A receptor antagonist, CR1409 and CCK-B antagonist, CR2945 were applied to sling and clasp fibers and their pK(B) values were calculated. Sling fibers produced significant contractions following exposure to CCK-8 and gastrin-17, while clasp fibers had less responses to the two agents. CR1409 and CR2945 inhibited responses of sling to CCK-8 in a concentration-dependent fashion. The inhibition effects of the two antagonists on clasp fibers were not measurable because there was a mild contraction of the fiber in response to CCK-8. The contractions generated by sling fibers following exposure to CCK and gastrin are greater than that produced by clasp fibers. CCK-A receptors are more important for the generation of contractions by the sling fibers, whereas both CCK-A and CCK-B receptors are involved in the functional regulation of the clasp fibers. [Corrections added after online publication 28 April 2008: in the Background and Aims section of the preceding abstract, all instances of 'CKK' were corrected to 'CCK'. In the final sentence of the abstract 'CCKA'was corrected to 'CCK-A'. In the article title '(CKK)' was corrected to '(CCK)'.].


Investigating the Morphology and Mechanics of Biogenic Hierarchical Materials at and below Micrometer Scale.
Investigating and understanding the intrinsic material properties of biogenic materials, which have evolved over millions of years into admirable structures with difficult to mimic hierarchical levels, holds the potential of replacing trial-and-error-based materials optimization in our efforts to make synthetic materials of similarly advanced complexity and properties. An excellent example is biogenic silica which is found in the exoskeleton of unicellular photosynthetic algae termed diatoms. Because of the complex micro- and nanostructures found in their exoskeleton, determining the intrinsic mechanical properties of biosilica in diatoms has only partly been accomplished. Here, a general method is presented in which a combination of in situ deformation tests inside an SEM with a realistic 3D model of the frustule of diatom Craspedostauros sp. (C. sp.) obtained by electron tomography, alongside finite element method (FEM) simulations, enables quantification of the Young's modulus (E = 2.3 ± 0.1 GPa) of this biogenic hierarchical silica. The workflow presented can be readily extended to other diatom species, biominerals, or even synthetic hierarchical materials.


Sensitive analysis of two barbiturates in human urine by capillary electrophoresis with sample stacking induced by moving reaction boundary.
An on-line stacking method based on moving reaction boundary (MRB) was developed for the sensitive determination of barbital and phenobarbital in human urine via capillary electrophoresis (CE). The optimized conditions for the method are: 60 mmol L(-1) pH 11.0 Gly-NaOH as the background electrolyte, 10 mmol L(-1) pH 5.5 Gly-HCl as sample buffer, secobarbital as the internal standard (IS), 12.5 kV, 1.4 psi 10s sample injection, 75 microm ID 60.2 cm total length (50 cm effective length) capillary and 214 nm detect wavelength. Under the optimized conditions, the method can well stack and separate barbital and phenobarbital in urine samples and result in 20.5-fold and 22.6-fold improvement in concentration sensitivity for barbital and phenobarbital, respectively. Furthermore, the method holds: (1) good linear calibration functions for the two target compounds (correlation coefficients r>0.999), (2) low limits of detection (0.27 microg mL(-1) for barbital and 0.26 microg mL(-1) for phenobarbital), (3) low limits of quantification (0.92 microg mL(-1) for barbital and 0.87 microg mL(-1) for phenobarbital), (4) good precision (R.S.D. of intra-day and inter-day less than 5.38% for barbital and 1.67% for phenobarbital, respectively) and (5) high recoveries at three concentration levels (90.27-106.36% for barbital and 93.05-113.60% for phenobarbital in urine). The method is simple, sensitive and efficient, and can fit to the need of clinical and forensic toxicology.


The Hydrating Effects of Hypertonic, Isotonic and Hypotonic Sports Drinks and Waters on Central Hydration During Continuous Exercise: A Systematic Meta-Analysis and Perspective.
Body-fluid loss during prolonged continuous exercise can impair cardiovascular function, harming performance. Delta percent plasma volume (dPV) represents the change in central and circulatory body-water volume and therefore hydration during exercise; however, the effect of carbohydrate-electrolyte drinks and water on the dPV response is unclear. To determine by meta-analysis the effects of ingested hypertonic (> 300 mOsmol kg-1), isotonic (275-300 mOsmol kg-1) and hypotonic (< 275 mOsmol kg-1) drinks containing carbohydrate and electrolyte ([Na+] < 50 mmol L-1), and non-carbohydrate drinks/water (< 40 mOsmol kg-1) on dPV during continuous exercise. A systematic review produced 28 qualifying studies and 68 drink treatment effects. Random-effects meta-analyses with repeated measures provided estimates of effects and probability of superiority (p+) during 0-180 min of exercise, adjusted for drink osmolality, ingestion rate, metabolic rate and a weakly informative Bayesian prior. Mean drink effects on dPV were: hypertonic - 7.4% [90% compatibility limits (CL) - 8.5, - 6.3], isotonic - 8.7% (90% CL - 10.1, - 7.4), hypotonic - 6.3% (90% CL - 7.4, - 5.3) and water - 7.5% (90% CL - 8.5, - 6.4). Posterior contrast estimates relative to the smallest important effect (dPV = 0.75%) were: hypertonic-isotonic 1.2% (90% CL - 0.1, 2.6; p+ = 0.74), hypotonic-isotonic 2.3% (90% CL 1.1, 3.5; p+ = 0.984), water-isotonic 1.3% (90% CL 0.0, 2.5; p+ = 0.76), hypotonic-hypertonic 1.1% (90% CL 0.1, 2.1; p+ = 0.71), hypertonic-water 0.1% (90% CL - 0.8, 1.0; p+ = 0.12) and hypotonic-water 1.1% (90% CL 0.1, 2.0; p+ = 0.72). Thus, hypotonic drinks were very likely superior to isotonic and likely superior to hypertonic and water. Metabolic rate, ingestion rate, carbohydrate characteristics and electrolyte concentration were generally substantial modifiers of dPV. Hypotonic carbohydrate-electrolyte drinks ingested continuously during exercise provide the greatest benefit to hydration.


Curcumin enhances cytotoxicity of chemotherapeutic agents in prostate cancer cells by inducing p21(WAF1/CIP1) and C/EBPbeta expressions and suppressing NF-kappaB activation.
The modulatory effects and molecular mechanisms of curcumin (CCM) on the cytotoxicity of chemotherapeutic agents to prostate cancer cells were explored. The combined effects of CCM and chemotherapeutic agents were examined by three different administration schedules (one concurrent and two sequential treatments) in two androgen-independent prostate cancer (AIPC) cells (PC-3 and DU145). Alteration of cell cycle progression, protein levels, and transcriptional activation in PC-3 cells were assayed by flow cytometry, Western blotting, and gel shift assay, respectively. The combined effects of CCM --> chemotherapeutic agent schedule showed the greatest synergistic cytotoxicity when compared to the other two schedules in both cells. CCM induced a significant G1 arrest in PC-3, which may be mediated by the induction of p21(WAF1/CIP1) and C/EBPbeta. Moreover, CCM was able to inhibit both the constitutional and TNF-alpha-induced NF-kappaB activation in a time-dependent manner. The incorporation of CCM into cytotoxic therapies may be a promising strategy for the treatment of AIPC.


Development of a seedling inoculation technique for rapid evaluation of soybean for resistance to Phomopsis longicolla under controlled conditions.
Phomopsis seed decay (PSD) of soybean (Glycine max L. Merr.) is caused primarily by the seed-borne fungal pathogen Phomopsis longicolla T. W. Hobbs. The PSD disease reduces seed quality and yield worldwide. Development of effective techniques to evaluate soybean for resistance to PSD can facilitate identification of new sources of host resistance to manage the disease. This study was undertaken to develop and utilize a rapid cut-seedling inoculation technique to evaluate soybean genotypes for resistance to P. longicolla under controlled conditions. There were no significant differences in stem lesion length determined as the area under disease progress curve at 24 °C and 30 °C. The 21 and 14-day-old seedlings were more susceptible than the older seedlings. Inoculation with 7 or 14-day-old pathogens caused higher values of AUDPC than older pathogen cultures. Isolates MS17-1 was the most aggressive isolate from the test of 25 isolates from seven states in the U.S. Eighteen previously reported field PSD-resistant accessions had significantly lower AUDPC than the susceptible checks and other entries (P ≤ 0.05). This study provided rapid evaluation of soybeans for reaction to P. longicolla and identification of PSD-resistant genotypes. Although PSD is a soybean seed disease, results from the cut-seedling inoculation assays without waiting a whole growing season were comparable to those obtained from field tests. Additionally, concerns about the environmental effects and uneven distribution of the pathogen in the field were ameliorated. The cut-seedling inoculation technique can also be used to speed up evaluation of PSD populations for the discovery of PSD-resistance gene(s), and high throughput phenotyping of seed diseases at seedling stage for genetics and genomic studies.


New Mechanisms and Targets of Subarachnoid Hemorrhage: A Focus on Mitochondria.
Spontaneous subarachnoid hemorrhage (SAH) accounts for 5-10% of all strokes and is a subtype of hemorrhagic stroke that places a heavy burden on health care. Despite great progress in surgical clipping and endovascular treatment for ruptured aneurysms, cerebral vasospasm (CVS) and delayed cerebral ischemia (DCI) threaten the long-term outcomes of patients with SAH. Moreover, there are limited drugs available to reduce the risk of DCI and adverse outcomes in SAH patients. New insight suggests that early brain injury (EBI), which occurs within 72 h after the onset of SAH, may lay the foundation for further DCI development and poor outcomes. The mechanisms of EBI mainly include excitotoxicity, oxidative stress, neuroinflammation, blood-brain barrier (BBB) destruction, and cellular death. Mitochondria are a double-membrane organelle, and they play an important role in energy production, cell growth, differentiation, apoptosis, and survival. Mitochondrial dysfunction, which can lead to mitochondrial membrane potential (Δψm) collapse, overproduction of reactive oxygen species (ROS), release of apoptogenic proteins, disorders of mitochondrial dynamics, and activation of mitochondria-related inflammation, is considered a novel mechanism of EBI related to DCI as well as post-SAH outcomes. In addition, mitophagy is activated after SAH. In this review, we discuss the latest perspectives on the role of mitochondria in EBI and DCI after SAH. We emphasize the potential of mitochondria as therapeutic targets and summarize the promising therapeutic strategies targeting mitochondria for SAH.


Inhibition of several protein phosphatases by a non-covalently interacting microcystin and a novel cyanobacterial peptide, nostocyclin.
Microcystins produced by cyanobacterial 'blooms' in reservoirs and lakes pose significant public health problems because they are highly toxic due to potent inhibition of protein serine/threonine phosphatases in the PPP family. A dehydrobutyrine (Dhb)-containing microcystin variant [Asp3, ADMAdda5, Dhb7]microcystin-HtyR isolated from Nostoc sp. was found to potently inhibit PP1, PP2A, PPP4 and PPP5 with IC50 values similar to those of microcystin-LR. However, in contrast to microcystin-LR, which forms a covalent bond with a cysteine residue in these protein phosphatases, Asp,ADMAdda,Dhb-microcystin-HtyR did not form any covalent interaction with PP2A. Since the LD50 for Asp,ADMAdda,Dhb-microcystin-HtyR was 100 microg kg(-1) compared to 50 microg kg(-1) for microcystin-LR, the data indicate that the non-covalent inhibition of protein phosphatases accounts for most of the harmful effects of microcystins in vivo. A 3-amino-6-hydroxy-2-piperidone containing cyclic peptide, nostocyclin, also isolated from Nostoc sp., was non-toxic and exhibited more than 500-fold less inhibitory potency towards PP1, PP2A, PPP4 and PPP5, consistent with the conclusion that potent inhibition of one or more these protein phosphatases underlies the toxicity of microcystins, both lacking and containing Dhb.


Relationship between the expression of CD44v6 and development, progress, invasion and metastasis of laryngeal carcinoma.
The expression of CD44v6 and its relationship with the development, progress, invasion and metastasis of laryngeal carcinoma was investigated. The expression and content of CD44v6 mRNA in tissuess were detected by both RT-PCR and FCM which were respectively extracted from normal laryngeal mucosa, leukoplakia of larynx, laryngeal papilloma, polyp of vocal cord, tissues of laryngeal carcinoma, metastatic and nonmetastatic lymph nodes of neck, and tissues close to carcinoma. The outcome of RT-PCR indicated that the expression rate of CD44v6 mRNA involved in tissues of laryngeal carcinoma and metastatic lymph nodes of neck was the highest (90%-100%) compared with that of leukoplakia of larynx, laryngeal papilloma, tissues close to carcinoma by 0.5 cm (55.56%-60.00%) and that of normal laryngeal mucosa, polyp of vocal cord, nonmetastatic lymph nodes and tissues close to carcinoma by 1.0 cm was the lowest (13.33%-20%). The result from FCM was highly consistent with that from RT-PCR. It was suggested that CD44v6 was closely related with the development, progress, invasion and metastasis of laryngeal carcinoma. The outcome from the tissues close to carcinoma by different distance could do help to the determination of incisal edge in surgery abstractly.


Cocaine-paired cues activate aversive representations in accumbens neurons.
Aversive states are proposed to drive addiction. Here, Wheeler and colleagues show that drug-associated cues come to activate neural representations of aversive information in nucleus accumbens and that this activation predicts subsequent drug use. These remarkable data identify a potential neural substrate through which aversive affective representations may motivate drug use.


Evaluation of sample extraction methods for proteomics analysis of green algae Chlorella vulgaris.
Many protein extraction methods have been developed for plant proteome analysis but information is limited on the optimal protein extraction method from algae species. This study evaluated four protein extraction methods, i.e. direct lysis buffer method, TCA-acetone method, phenol method, and phenol/TCA-acetone method, using green algae Chlorella vulgaris for proteome analysis. The data presented showed that phenol/TCA-acetone method was superior to the other three tested methods with regards to shotgun proteomics. Proteins identified using shotgun proteomics were validated using sequential window acquisition of all theoretical fragment-ion spectra (SWATH) technique. Additionally, SWATH provides protein quantitation information from different methods and protein abundance using different protein extraction methods was evaluated. These results highlight the importance of green algae protein extraction method for subsequent MS analysis and identification.


Host response in bovine mastitis experimentally induced with Staphylococcus chromogenes.
An experimental infection model was developed to study host response to intramammary infection in cows caused by Staphylococcus chromogenes. CNS intramammary infections have become very common in modern dairy herds, and they can remain persistent in the mammary gland. More information would be needed about the pathophysiology of CNS mastitis, and an experimental mastitis model is a means for this research. Six primiparous Holstein-Friesian cows were challenged with S. chromogenes 4 weeks after calving. One udder quarter of each cow was inoculated with 2.1 x 10(6)cfu of S. chromogenes. All cows became infected and clinical signs were mild. Milk production of the challenged quarter decreased on average by 16.3% during 7 days post-challenge. Cows eliminated bacteria in a few days, except for one cow which developed persistent mastitis. Milk indicators of inflammation, SCC and N-acetyl-beta-D-glucosaminidase (NAGase) returned to normal within a week. Milk NAGase activity increased moderately, which reflects minor tissue damage in the udder. Concentrations of serum amyloid A (SAA) and milk amyloid A (MAA) were both elevated at 12h PC. MAA was affected by the milking times, and was at its highest before the morning milking. In our experimental model, systemic acute phase protein response with SAA occurred as an on-off type reaction. In conclusion, this experimental model could be used to study host response in CNS mastitis caused by the main CNS species and also for comparison of the host response in a mild intramammary infection and in more severe mastitis models.


Laminin therapy for the promotion of muscle regeneration.
Muscle regeneration is essentially due to activation of satellite cells, which can be isolated and amplified ex vivo, thus representing good candidates for cell therapy. Accumulating data show that the local microenvironment plays a major role during muscle regeneration. In the satellite cell niche, a major extracellular matrix protein is laminin. Human myoblasts transplanted into immunodeficient mice are preferentially located in laminin-enriched areas. Additionally, laminin-111 enhances myoblast proliferation in vitro and increases expression of the α7β1 integrin-type laminin receptor. Intramuscular injection of laminin-111 ameliorates muscular pathology in mdx mice, protecting muscle fibers from damage. Moreover, transplantation of human myoblasts with laminin-111 into Rag/mdx immunodeficient recipients improved efficacy of myoblast transplantation, increasing the number of human dystrophin-positive myofibres. Taken together, these data strongly indicate that exogenous laminin can ameliorate the regeneration process in different models of muscular dystrophies and can be instrumental for improving cell therapy aiming at repairing the degeneration/regeneration process in skeletal muscle.


Action of mannitol in various immunological experimental models.
The authors describe the inhibiting action of mannitol after repeated administration of low subcutaneous doses in a number of experimental immunological models. For example, in the rat it produces a reduction of the secondary arthritis of Freund's adjuvant polyarthritis and also of the pleurisy due to Bordetella pertussis hypersensitivity. In the mouse it reduces the reaction of delayed hypersensitivity to sheep red cells. Its action is also marked against ovalbumin-induced active skin anaphylaxis in the albino guinea-pig and on IgE synthesis in the rat. Moreover, after several injections it produces a reduction of carbon phagocytosis in the mouse. At the doses at which the effect appeared, no action could be found on various models of acute non-immune inflammation, diuresis, blood pressure, hematocrit and protein and plasma sodium levels.


In silico evidence of beauvericin antiviral activity against SARS-CoV-2.
Scientists are still battling severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for the coronavirus 2019 (COVID-19) pandemic so human lives can be saved worldwide. Secondary fungal metabolites are of intense interest due to their broad range of pharmaceutical properties. Beauvericin (BEA) is a secondary metabolite produced by the fungus Beauveria bassiana. Although promising anti-viral activity has previously been reported for BEA, studies investigating its therapeutic potential are limited. The objective of this study was to assess the potential usage of BEA as an anti-viral molecule via protein-protein docking approaches using MolSoft. In-silico results revealed relatively favorable binding energies for BEA to different viral proteins implicated in the vital life stages of this virus. Of particular interest is the capability of BEA to dock to both the main coronavirus protease (Pockets A and B) and spike proteins. These results were validated by molecular dynamic simulation (Gromacs). Several parameters, such as root-mean-square deviation/fluctuation, the radius of gyration, H-bonding, and free binding energy were analyzed. Computational analyses revealed that interaction of BEA with the main protease pockets in addition to the spike glycoprotein remained stable. Altogether, our results suggest that BEA might be considered as a potential competitive and allosteric agonist inhibitor with therapeutic options for treating COVID-19 pending in vitro and in vivo validation.


Low dietary riboflavin but not folate predicts increased fracture risk in postmenopausal women homozygous for the MTHFR 677 T allele.
The MTHFR C677T polymorphism is associated with mildly elevated homocysteine levels when folate and/or riboflavin status is low. Furthermore, a mildly elevated homocysteine level is a risk factor for osteoporotic fractures. We studied whether dietary intake of riboflavin and folate modifies the effects of the MTHFR C677T variant on fracture risk in 5,035 men and women from the Rotterdam Study. We found that the MTHFR C677T variant interacts with dietary riboflavin intake to influence fracture risk in women. The MTHFR C677T polymorphism is associated with mildly elevated homocysteine (Hcy) levels in the presence of low folate and/or riboflavin status. A mildly elevated Hcy level was recently identified as a modifiable risk factor for osteoporotic fracture. We studied whether dietary intake of riboflavin and folate modifies the effects of the MTHFR C677T polymorphism on BMD and fracture risk. We studied 5,035 individuals from the Rotterdam Study, >or=55 yr of age, who had data available on MTHFR, nutrient intake, and fracture risk. We performed analysis on Hcy levels in a total of 666 individuals, whereas BMD data were present for 4,646 individuals (2,692 women). In the total population, neither the MTHFR C677T polymorphism nor low riboflavin intake was associated with fracture risk and BMD. However, in the lowest quartile of riboflavin intake, female 677-T homozygotes had a 1.8 (95% CI: 1.1-2.9, p = 0.01) times higher risk for incident osteoporotic fractures and a 2.6 (95% CI: 1.3-5.1, p = 0.01) times higher risk for fragility fractures compared with the 677-CC genotype (interaction, p = 0.0002). This effect was not seen for baseline BMD in both men and women. No significant influence was found for dietary folate intake on the association between the MTHFR C677T genotype and fracture risk or BMD. In the lowest quartile of dietary riboflavin intake, T-homozygous individuals (men and women combined) had higher (22.5%) Hcy levels compared with C-homozygotes (mean difference = 3.44 microM, p = 0. 01; trend, p = 0.02). In this cohort of elderly whites, the MTHFR C677T variant interacts with dietary riboflavin intake to influence fracture risk in women.


Prevention and control of OQDS (olive quick decline syndrome) outbreaks caused by Xylella fastidiosa.
In Southern Italy, since 2013, there has been an ongoing Olive Quick Decline Syndrome (OQDS) outbreak, due to the bacterium Xylella fastidiosa, which has caused a dramatic impact from both socio-economic and environmental points of view. The main players involved in OQDS are represented by the insect vector, Philaenus spumarius, its host plants (olive trees and weeds) and the bacterium, X. fastidiosa. Current agronomic practices are mainly based on uprooting the sick olive trees and their surrounding ones, with later installment of olive cultivars more resistant to the bacterium infection. Unfortunately, both of these practices are having an undesirable impact on the environment (most of these olive trees were monumental ones) and on the economy. Based on a mathematical model expressed in terms of a nontrivial system of ordinary differential equations, our analysis has provided a clear picture of all possible steady states (feasible equilibria) and their stability properties, corresponding to a variety of different parameter scenarios; all of this has been illustrated by a set of computational experiments. A significant original contribution of this paper is the proof of the global asymptotic stability of each of the feasible equilibria under its existence assumptions, a fact that excludes multiple equilibria under the given conditions. It has emerged that the removal of a suitable amount of weed biomass (host plants of the juvenile stages of the insect vector of X. fastidiosa) from olive orchards and surrounding areas leads to the eradication of the epidemic, without requiring neither the removal nor the substitution of the existing olive trees.


Gene Therapy for the Treatment of Neurological Disorders: Metabolic Disorders.
Metabolic disorders comprise a large group of heterogeneous diseases ranging from very prevalent diseases such as diabetes mellitus to rare genetic disorders like Canavan Disease. Whether either of these diseases is amendable by gene therapy depends to a large degree on the knowledge of their pathomechanism, availability of the therapeutic gene, vector selection, and availability of suitable animal models. In this book chapter, we review three metabolic disorders of the central nervous system (CNS; Canavan Disease, Niemann-Pick disease and Phenylketonuria) to give examples for primary and secondary metabolic disorders of the brain and the attempts that have been made to use adeno-associated virus (AAV) based gene therapy for treatment. Finally, we highlight commonalities and obstacles in the development of gene therapy for metabolic disorders of the CNS exemplified by those three diseases.


Dynamic MR imaging analysis of instability in the injured Lisfranc joint with an MRI-compatible foot stressor device.
To evaluate the applicability of an MRI-compatible foot stressor device in patients with image-proven or clinically suspected Lisfranc joint injuries. This prospective study evaluated Lisfranc joint injury by utilizing a joint-specific, stress device that was engineered to replicate weightbearing and physical examination maneuvers. Sixteen patients with either clinically suspected or image-proven Lisfranc joint injuries were recruited from September 2018 to November 2019 (9 men, 7 women; mean age, 39.3 years; age range, 14-68 years). Resting and stressed MR sequences of the injured and non-injured feet were obtained. Measured values for Lisfranc interval widths, dorsal tarsometatarsal subluxations, and lambda-angles were subtracted between the stressed and resting images to calculate net stress-induced changes. A graded injury schema was used to measure significance. The foot stressor device reliably generated stress-induced changes in the Lisfranc joint during dynamic MRI examination. All morphologically abnormal ligaments on resting images demonstrated stress-induced changes, whereas all morphologically normal ligaments lacked evidence of instability. More severely injured Lisfranc ligaments allowed greater Lisfranc joint widening (IOL, p < 0.001; PCL, p < 0.001; DCL, p < 0.001). More highly graded DCL injuries allowed greater dorsal TMT subluxation when present (p < 0.001). Angular gain in the midfoot (lambda-angle) correlated with the graded IOL score (p < 0.001). Acute-to-subacute injuries demonstrated greater inducible changes than chronic injuries (p = 0.047). Seven patients underwent surgery and nine patients received physical therapy. Stress-induced changes in the midfoot provided information on the degree of ligament pathology and associated joint instability in Lisfranc joint injuries.


Grey scale ultrasonic examination of the thyroid gland.
High resolution grey scale echography is an accurate, non-invasive means of investigating thyroid lesions. The structural information which it provides complements that of the functional status of the gland obtained from radionuclide scanning. The principles and practice of the technique are discussed and clinical examples are presented.


Leveraging COVID-19 vaccination to promote hepatitis C screening.
Health care initiatives, such as hepatitis C virus (HCV) screening, have been greatly overshadowed by the corona virus disease 2019 (COVID-19) pandemic. However, COVID-19 vaccination programs also provide an opportunity to engage with a high volume of people in a health care setting. We collaborated with a large COVID vaccination center to offer HCV point-of-care testing followed by dried blood spot collection for HCV RNA. Additionally, this opportunity was used to evaluate the practical significance of a 5-minute version of the OraQuick HCV antibody test in lieu of the standard 20-minute test. We tested 2317 individuals; 31 were HCV antibody positive and six were RNA positive of which four were treated and reached sustained virological response. Over a third of those surveyed said they would not have participated had the test required 20 minutes. Conclusion : Colocalizing HCV testing and linkage to care at a COVID vaccination clinic was found to be highly feasible; furthermore, a shortened antibody test greatly improves the acceptance of testing.


Neisseria gonorrhoeae prevalence in females in São Paulo, Brazil: surveillance of the infection over a 11-year period.
The prevalence of infection by Neisseria gonorrhoeae is the second highest among bacterial sexually transmitted infections worldwide. It can lead to severe complications, especially in the female reproductive system. So, the objective of the present study was to assess the prevalence of Neisseria gonorrhoeae infection in a large population of female patients from a private health service in São Paulo (Brazil), thereby identifying the main age groups affected by the agent and how prevalence progressed over time. A cross-sectional study was conducted based on the results of all molecular biology tests performed for detecting Neisseria gonorrhoeae. The tests were conducted between January 2005 and December 2015. Positive test results were grouped by year and age group. Of those tests, 35,886 were considered eligible for the statistical data. The overall prevalence of Neisseria gonorrhoeae infection in the study population was 0.4%. A higher prevalence of infection was observed in the group whose participants' age was ≤ 25 years, at a rate of 0.6%. There was no significant increase or decrease in the number of positive test results over time. The prevalence of the infection in those aged between 10 and 19 years, 20-29 years, 30-39 years, 40-49 years, 50-59 years, and 60 years or above was respectively 0.87%, 0.50%, 0.36%, 0.22%, 0.17%, and 0.26%. The screening of asymptomatic young women would have the potential to reduce infection, transmission, and sequelae of infection by this agent.


Efficient production of recombinant human transcription factor IIE.
Transcription factor IIE (TFIIE) is a general initiation and promoter escape factor for RNA polymerase II composed of p56 (TFIIE-alpha) and p34 (TFIIE-beta) subunits. Our laboratories experienced difficulty producing adequate quantities of recombinant human TFIIE-alpha for in vitro studies using available clones. We therefore re-engineered the TFIIE subunit production vectors and tested various Escherichia coli host strains to optimize expression. We report a much-improved system for production of pure, soluble, and active TFIIE complex for in vitro studies.


Significance of portosystemic shunt on graft survival in liver transplantation: a rat model.
Distal splenorenal shunt effectively controls bleeding from esophageal and gastric varices but has a different effect on liver transplantation. This study sought to develop an animal model in rats to mimic the recipient with a portosystemic shunt and to investigate its hemodynamic consequences on liver transplantation. We prepared 5 groups of allogeneic or syngeneic rat liver transplantation models with versus without portosystemic shunt, to investigate its effects on graft survival and portal flow. To explore the effects of excessive portal flow on graft survival in small-for-size liver transplantation, we transplanted partial liver grafts into syngeneic recipients. In allogeneic combinations, graft survival among the shunt group was shortened compared with their control counterparts. The graft survival of the large shunt group was significantly lower than that of a small shunt or without shunt group in a syngeneic liver transplantation model. Portal blood pressure of the large shunt group was significantly lower than that of the small shunt group. In contrast, excessive portal flow resulted in dysfunction of liver graft in small-for-size liver transplantation. These results suggested that reduction in portal flow by portosystemic shunt lead to an acceleration of acute rejection and subsequent liver graft dysfunction, but it may be applicable to regulate the excessive portal flow in small-for-size transplantations. This study showed a valuable model mimicking the recipient with a portosystemic shunt.


Golgi matrix protein gene, Golga3/Mea2, rearranged and re-expressed in pachytene spermatocytes restores spermatogenesis in the mouse.
In a transgenic mouse, Golga3/Mea2 gene (human homolog: GOLGA3/golgin-160) was disrupted by a translocation at the site of the transgene integration. Exons 8-24 of the disrupted gene remained intact and formed a fusion gene (DeltaMea2) with the antisense strand of E. coli-derived transgene by means of a cryptic splice signal in there. The protein product of DeltaMea2, virtually a form truncated to 2/3 of the normal size, localized to Golgi apparatus of pachytene spermatocytes and round spermatids. DeltaMea2 expression was specific to the testis, but varied among separate seminiferous tubules. It also showed variation among homozygous individuals from 0.5 to 4.3% of the wild type (wt) level. At the lowest levels, neither spermatids nor spermatozoa were present in the homozygous testes, but when the expression of DeltaMea2 increased to 4.3% of the wt level, high sperm production was restored and a sporadic (1/22) fertile homozygous male was obtained. The earliest apoptotic degeneration of pachytene spermatocytes evidenced at 17 dpp in homozygous testes in some discrete seminiferous tubules was preceded by DeltaMea2 expression in a variegated fashion at 16 dpp. These results consistently indicated that in homozygous testes, the pachytene spermatocytes which failed to express DeltaMea2 may undergo apoptotic degeneration. Golga3/Mea2, and DeltaMea2 in homozygotes, in a certain excessive amount may be important for survival of pachytene spermatocytes in the mouse.


Techniques for analysing and monitoring during continuous bio-hydrogenation of kerosene from palm oils.
In this research work, analytical, experimental methods and monitoring techniques of bio-hydrogenated kerosene (BHK) production in continuous mode were presented. Two kinds of raw materials obtained from palm processing plant named as refined bleached deodorised palm oil (RPO) and palm kernel oil (PKO) were converted into BHK via hydrocracking reaction catalysed by Pd/Al2O3 catalyst in pilot scale. Firstly, both of RPO and PKO were pretreated by thermal technique. Subsequently, fatty acid compositions of palm oils were analysed by Gas Chromatography (GC). Then, hydrocracking reaction of RPO and PKO were separately conducted in continuous high pressure packed bed reactor (HPPBR). After reaction, crude-biofuel was refined into BHK via fractional distillation. In addition, some properties of BHK obtained from RPO/PKO such as were C, H, O elements, freezing point, flash points, total acid number and carbon distribution were analysed following the ASTM and UOP 915 standards.•Thermal pretreatment of refined bleached deoderised palm oil (RPO) and palm kernel oil (PKO).•Continuous hydrocracking reaction of palm oil was conducted in pilot scale.•Characterisation of bio-hydrogenated kerosene obtained from palm oil.


The effect of statistical sampling error on reliability and validity.
The essential purpose of sampling is to select a proportion, a sample, of the members of a group, a statistical population, and to do so in such a way that any data obtained from the sample can be used to describe the statistical population in a meaningful way. A description of a statistical population is needed when it has become necessary to resolve a problem that affects that population. It shows the severity of the problem, the extent of the problem, and the effectiveness of the action taken to resolve the problem. The statistical population is described by first describing the sample, and then converting this description into a description of the statistical population.


Lymphovascular space invasion in uterine corpus cancer: What is its prognostic significance in the absence of lymph node metastases?
Lymphovascular space invasion (LVSI) is a poor prognostic indicator in uterine cancer, primarily due to its association with lymph node metastases. We sought to determine if LVSI provides any prognostic information for uterine cancer subjects in the absence of nodal disease. A retrospective review was performed using a database of women treated for uterine cancer at MUSC from 2005 to 2012. Subjects with negative nodes after complete staging were identified. Multiple regression modeling was used to adjust for demographic and histopathologic covariates. The C-index was calculated for models of survival that included LVSI and those that did not. Competing risks analysis was conducted to examine factors associated with time to recurrence. Two hundred and five subjects were completely staged and had negative nodes, 24 with LVSI and 181 without. Factors significantly associated with survival included age, race, stage, grade, histology, and LVSI. Regression models for recurrence-free survival (RFS) and overall survival (OS) had similar C-indices regardless of whether LVSI was included. Competing risks analysis confirmed no significant difference in time to recurrence for subjects with LVSI compared to those without, after adjusting for other prognostic factors (P=0.53). LVSI is associated with shorter recurrence-free and overall survival in uterine cancer subjects with negative lymph nodes. However, after adjusting for other prognostic factors, LVSI status does not provide additional prognostic information. This finding suggests that recurrence-free and overall survival for uterine cancer patients with negative lymph nodes can be estimated without factoring in LVSI.


Psychosocial risk factors and mortality: a prospective study with special focus on social support, social participation, and locus of control in Norway.
The objective is to investigate the effect on mortality of psychosocial variables, with special focus on social support, social participation, and locus of control. The study is designed as a prospective study with a 17 year follow up period, using univariate and multivariate proportional hazards regression analysis to estimate the predictive power of psychosocial variables, when controlling for sociodemographic and biological factors. The study is based on a population sample randomly drawn from different neighbourhoods of Oslo in 1975/76, for the purpose of surveying health, in particular mental health, in relation to various social and psychosocial variables. The initial data were gathered by structured interviewing, whereas the data about mortality and cause of death, was gathered from the Central Bureau of Statistics. The initial sample included 1010 persons above the age of 18 years, with no upper age limit. The follow up with respect to mortality covered the whole sample, with the exception of a very few who had left the country. When controlling for socio-demographic and biological factors, low social participation, and to a lesser extent, few close relationships and external locus of control, were associated with increased mortality. The effect of social participation and locus of control may indicate that life style, and individual psychological resources, are at least as important for survival as support from others in stressful life situations.


Using delta/DRG diagrams and decision tree analysis to select a cost-effective surgery for cholecystitis.
Many studies have attempted cost analysis of laparoscopic cholecystectomy as compared to open cholecystectomy. However, these analyses have included costs, charges, expenses, etc., and at times they have been used interchangeably. This paper demonstrates how DRG diagrams containing charges and length-of-stay, preoperative prediction of conversion rates, decision-tree construction and sensitivity analysis can be used to select the most cost-efficient operation for a given patient with cholecystitis. A Delta DRG analysis for complicated cholecystectomy (DRG 195) showed the hospital to be an extreme outlier in both charges and length of stay. Record review indicated that 55% of the cases were converted laparoscopic cholecystectomies and the remainder were aged or younger patients with advanced disease. Chart and literature review determined the causes and the probability of conversion. Data were then placed into decision-tree and sensitivity analyses. The most cost-effective operation for a given probability of conversion was demonstrated. Three preoperative findings and combinations of each predicted conversion rates and analysis showed that the charge of laparoscopic cholecystectomy must be held below the range of $5,361-$13,084 to make routine laparoscopic cholecystectomy cost-effective. This method demonstrated that using Delta/DRG, decision-tree and sensitivity analysis offers physicians, hospitals and other health-care providers a method of evaluating the treatment of DRG categories to determine the most cost-effective management.


Fluorescence imaging of blood-brain barrier disruption.
Pathological alterations of the blood-brain barrier (BBB) can be topographically heterogeneous. The goal of this study was to develop a method to assess rapidly the magnitude and spatial distribution of permeability changes. Rats were perfused via the common carotid arteries with Ringer's solution containing sodium fluorescein (NF) and Evans Blue albumin (EB). Global NF uptake was determined by fluorimetry and EB uptake was determined by absorbance spectroscopy. NF uptake was linear in control animals and at a rate comparable to sucrose, whereas uptake of EB was negligible. Infusion of 1.6 M mannitol immediately prior to perfusion significantly increased uptake of NF while EB uptake was unchanged. BBB disruption was confirmed by confocal microscopy of fresh-frozen sections. In control animals, NF and EB staining were limited to the edges of slices and to the circumventricular organs. In mannitol-treated animals, heavy NF staining was observed throughout the brain, and EB staining was localized around some microvessels. In animals given a approximately 500 microl air embolus prior to perfusion, a discrete area of NF and EB staining could be observed near the ventral midline, while the rest of the brain remained unaltered. We find that brain perfusion with NF/EB enables a rapid, reliable, and highly sensitive assessment of global BBB permeability and microscopic visualization of discrete BBB disruptions.


Activation of TLR4 induces severe acute pancreatitis-associated spleen injury via ROS-disrupted mitophagy pathway.
Severe acute pancreatitis (SAP) is complicated by systemic inflammatory response syndrome and multiple organ dysfunction, the disease will eventually result in death in almost half of the case. The spleen, as the largest immune organ adjacent to the pancreas, is prone to damage in SAP, thereby aggravating the damage of other organs and increasing mortality. However, to date, the research on the mechanism and treatment of spleen injury caused by SAP is still in its infancy. Herein, we investigated the mechanism of spleen injury, and explored the application potential of tuftsin for relieving spleen damage in SAP mice. Firstly, SAP mice model was constructed via the retrograde infusion of 3.5 % sodium taurocholate into the biliopancreatic duct. Then, we proved that the up-regulation of Toll-like receptor 4 (TLR4) in spleen would lead to the accumulation of reactive oxygen species (ROS) and mitochondrial dysfunction under SAP conditions. The splenic ROS and mitochondrial dysfunction could be improved by N-acetylcysteine (NAC) treatment or knocking out TLR4 in SAP mice. Meanwhile, we found that NAC treatment could also improve the autophagy of spleen tissue, suggesting that splenic ROS may affect impaired autophagy, causing the accumulation of damaged mitochondria, aggravating spleen damage. Furthermore, we verified the mechanism of spleen injury is caused by splenic ROS affecting PI3K/p-AKT/mTOR pathway-mediated autophagy. In addition, we detected the spleen injury caused by SAP could decrease the concentration of tuftsin in the serum of mice. Whereas, exogenous supplementation of tuftsin ameliorated the pathological damage, ROS accumulation, impaired autophagy, inflammation expression and apoptosis in damaged spleen. In summary, we verified the new mechanism of SAP-caused spleen damage that TLR4-induced ROS provoked mitophagy impairment and mitochondrial dysfunction in spleen via PI3K/p-AKT mTOR signaling, and the application potential of tuftsin in treating spleen injury, which might expand novel ideas and methods for the treatment of pancreatitis.


Gastric secretion during anaesthesia and its inhibition by metiamide and other drugs.
The histamine H2-receptor antagonist, metiamide, inhibits the acid gastric secretion produced by chloralose-urethane anaesthesia in dogs carrying gastric cannulae chronically. This secretion is also prevented by atropine and hexamethonium. "Spontaneous" gastric secretion of vagal origin in conscious dogs is also blocked by metiamide.


Low-frequency stimulation in anterior nucleus of thalamus alleviates kainate-induced chronic epilepsy and modulates the hippocampal EEG rhythm.
High-frequency stimulation (HFS) of the anterior nucleus of thalamus (ANT) is a new and alternative option for the treatment of intractable epilepsy. However, the responder rate is relatively low. The present study was designed to determine the effect of low-frequency stimulation (LFS) in ANT on chronic spontaneous recurrent seizures and related pathological pattern in intra-hippocampal kainate mouse model. We found that LFS (1 Hz, 100 μs, 300 μA), but not HFS (100 Hz, 100 μs, 30 μA), in bilateral ANT significantly decreased the frequency of spontaneous recurrent seizures, either non-convulsive focal seizures or tonic-clonic generalized seizures. The anti-epileptic effect persisted for one week after LFS cessation, which manifested as a long-term inhibition of the frequency of seizures with short (20-60 s) and intermediate duration (60-120 s). Meanwhile, LFS decreased the frequency of high-frequency oscillations (HFOs) and interictal spikes, two indicators of seizure severity, whereas HFS increased the HFO frequency. Furthermore, LFS decreased the power of the delta band and increased the power of the gamma band of hippocampal background EEG. In addition, LFS, but not HFS, improved the performance of chronic epileptic mice in objection-location task, novel objection recognition and freezing test. These results provide the first evidence that LFS in ANT alleviates kainate-induced chronic epilepsy and cognitive impairment, which may be related to the modulation of the hippocampal EEG rhythm. This may be of great therapeutic significance for clinical treatment of epilepsy with deep brain stimulation.


Tackling inequalities in health: experience in the UK.
During the past year, there has been increasing emphasis on inequalities in health, following the publication of policy documents on public health and health services by the new Government. Each District has to produce a Health Improvement Programme, which involves local authorities, voluntary agencies and businesses. Local targets for inequality may be required. The framework proposed by Government covers three levels: 1. Increase disability-free life expectancy/reduce inequalities 2. Healthy schools, for children/healthy workplace, for adults/healthy localities for older people 3. Heart disease and stroke/cancers/accidents/mental health. The text provides specific examples of existing projects for children and the role of the NHS as the biggest employer in UK.


A controlled study of the effect of sulfinpyrazone on platelet survival and on platelet-bound 14C-serotonin release in patients with previous myocardial infarction.
Preliminary data obtained in the ambit of Anturan Reinfarction Italian Study (ARIS) show that, in postmyocardial infarction, reduced platelet survival time occurred in hyperbetalipoproteinemic patients treated with placebo, but not in the group of patients treated with sulfinpyrazone (interaction between treatment and lipemic level is at p approximately equal to 0.1). The sulfinpyrazone effect on platelet survival is probably related to the release reaction inhibition as suggested by our ex vivo results on platelet-bound 14C-serotonin release.


Decreased tolerance to ethanol-induced hypothermia in long-term castrate male rats.
A potential role for central stores of vasopressin in the development of tolerance was studied in the long-term castrate rat. Vasopressin stores in the septal region are known to be dramatically depressed following long-term castration. Sprague-Dawley male rat littermates were castrated at 26 days of age or given a sham surgery. Experiments began when animals reached 130 days of age. Tolerance to the hypothermic effects of ethanol occurred in intact but not castrate animals over the course of six daily IP injections of 3.0 g/kg ethanol. Both groups exhibited tolerance to the length of time needed to return to baseline temperature over the 6 days of ethanol injections. Tolerance to this effect of ethanol was still evident in intact animals but not castrates following another injection of ethanol 1 week later. No tolerance developed to the rebound hyperthermia that occurred in both groups. Blood ethanol levels did not differ significantly between castrate and intact littermates administered a single dose of ethanol. Overall, these results support the hypothesis that endogenous vasopressin is involved in the development of some aspects of tolerance to ethanol.


Gastroprotective and toxicological evaluation of the Lithothamnion calcareum algae.
Lithothamnion calcareum is a red alga of the Corallinacea family whose main feature is the formation of calcium carbonate precipitate in its cell walls. L. calcareum is marketed as a nutritional supplement for calcium and other minerals in Brazil and other countries under the pharmaceutical name of Vitality 50+. In this study, gastroprotective and pre-clinical toxicity assays were performed on this product. Doses of 30, 120 and 480 mg/kg were used in the gastroprotective study on Wistar rats. A dose of 2000 mg/kg was used in the preclinical acute toxicity study and oral doses of 1000 and 2000 mg/kg were used in the subchronic toxicity evaluation. L. calcareum played no significant role in the protection of the rats' gastric mucosa, nor did it cause increase in gastric irritation. No impact on the acute toxicity test was identified. In the subchronic toxicity test, serum levels of albumin, total protein and calcium decreased, and creatinine levels increased, suggesting hypercalcemia and possible kidney damage associated with liver damage, given that the majority of these parameters were irreversible. Thus, this work aims to discuss the relationship of the high concentration of calcium in the product with the observed effects.


Fifty Years of a Dispute. A Triptych: Why Woodward?
This publication discusses R. B. Woodward's motivations, personality qualities, and chemical experiences that placed him in a perfect position to discover, along with Roald Hoffmann, the solution to the pericyclic no-mechanism problem, that is, to discover the Woodward-Hoffmann rules or, as Woodward preferred, the Principle of Conservation of Orbital Symmetry.


Binding of huperzine A and galanthamine to acetylcholinesterase, based on ONIOM method.
Binding energy calculations of huperzine A (HUP A) and galanthamine (GAL) to the binding pocket of the acetylcholinesterase enzyme (AChE) were studied. It was found that hydrogen bond formation and particular hydrogen π interactions exhibit the most significant contributions to the binding interaction of HUP A with Trp84 (W84) and Tyr130 (Y130), whereas no hydrogen bond was detected with Y130 of GAL binding. The interaction energies, calculated at the MP2 level between drugs and residues, demonstrate that the attractive interactions between GAL and residues at positions 84 and 130 were less than those for HUP A by 1.6 and 7.7 kcal·mol(-1), respectively. In addition, ONIOM3 results show that the binding energies of HUP A per pocket (-28.4 kcal mol(-1)) are higher than for GAL per pocket (-17.0 kcal·mol(-1)). The detailed understanding of these interactions can be useful for the design of specific inhibitors for the AChE binding site. The more efficient and specific inhibition of acetylcholinesterase may provide an enhanced treatment strategy in Alzheimer's disease compared to the currently available inhibitors. This study discusses interactions of the enzyme binding site with two ligands. The results may pave the way to the development of more potent inhibitors.


Machine Learning Approach for the Determination of the Best Cut-Off Points for Ki67 Proliferation Index in Adjuvant and Neo-Adjuvant Therapy Breast Cancer Patients.
This study aims to evaluate Ki67 cut-off points for differentiating low and high-risk patients based on survival and recurrence and find the best Ki67 cut-off points in breast cancer patients undergoing adjuvant and neoadjuvant therapy using machine learning methods. Patients with breast cancer treated at 2 referral hospitals between December 2000 and March 2021 who had invasive breast cancer entered this study. There were 257 patients in the neoadjuvant group and 2139 in the adjuvant group. A decision tree method was used to predict the likelihood of survival and recurrence. The 2-ensemble technique of RUSboost and bagged tree were imposed on the decision tree method to increase the accuracy of the determination. 80 percent of the data was used to train and validate the model, and 20% was used as a test. In adjuvant therapy breast cancer patients with Invasive ductal carcinoma (IDC) and Invasive lobular carcinoma (ILC) the cutoff points for survival were 20 and 10, respectively. For luminal A, luminal B, Her2 neu, and triple-negative adjuvant therapy patients' the cutoff points for survival were 25, 15, 20, and 20, respectively. For neoadjuvant therapy luminal A and luminal B group, survival cutoff points were 25 and 20, respectively. Despite variability in measurement and cut-off points, the Ki-67 proliferation index is still helpful in the clinic. Further investigation is needed to determine the best cut-off points for different patients. The sensitivity and specificity of Ki-67 cutoff point prediction models in this study could further prove its significance as a prognostic factor.


Suberoylanilide hydroxamic acid (SAHA) overcomes multidrug resistance and induces cell death in P-glycoprotein-expressing cells.
Multidrug resistance (MDR) mediated by the ATP-dependent efflux protein P-glycoprotein (P-gp) is a major obstacle to the successful treatment of many cancers. In addition to effluxing toxins, P-gp has been shown to protect tumor cells against caspase-dependent apoptosis mediated by Fas and tumor necrosis factor receptor (TNFR) ligation, serum starvation and ultraviolet (UV) irradiation. However, P-gp does not protect against caspase-independent cell death mediated by granzyme B or pore-forming proteins (perforin, pneumolysin and activated complement). We examined the effects of the chemotherapeutic hybrid polar compound suberoylanilide hydroxamic acid (SAHA) on P-gp-expressing MDR human tumor cell lines. In the CEM T-cell line, SAHA, a histone deacetylase inhibitor, induced equivalent death in P-gp-positive cells compared with P-gp-negative cells. Cell death was marked by the caspase-independent release of cytochrome c, reactive oxygen species (ROS) production and Bid cleavage that was not affected by P-gp expression. However, consistent with our previous findings, SAHA-induced caspase activation was inhibited in P-gp-expressing cells. These data provide evidence that P-gp inhibits caspase activation after chemotherapeutic drug treatment and demonstrates that SAHA may be of value for the treatment of P-gp-expressing MDR cancers.


Isolation and structure elucidation of a new prenylcoumarin from Murraya paniculata var. omphalocarpa (Rutaceae).
A new C-8 prenylated 5,7-dimethoxycoumarin named omphamurrayin was isolated from the leaves of Murraya paniculata var. omphalocarpa, and its structure was established as 5,7-dimethoxy-8-(1-oxo-2-senecioyl-3-methyl-3-butenyl)-2H-1-benzopyran-2-one on the basis of the spectroscopic evidence. The taxonomic status of M. paniculata var. omphalocarpa is briefly discussed, along with its synonymity to M. paniculata from the chemosystematic viewpoint.


Increase in endothelin B receptor expression in optic nerve astrocytes in endothelin-1 induced chronic experimental optic neuropathy.
The purpose of this study was to determine whether endothelin B (ETB) receptor levels in the optic nerve are related to retinal ganglion cell (RGC) loss in a model of chronic endothelin-1 (ET-1) induced optic neuropathy. RGCs of adult Brown Norway rats were first retrogradely labeled with fluorochrome from the superior colliculi. An osmotic minipump was surgically implanted 7 days later to deliver 10(-11) M (n = 9), 10(-9) M (n = 12) or 10(-7) M (n = 9) ET-1 to the retrobulbar optic nerve for 28 days. RGC survival was expressed as the ratio of RGC counts in experimental versus control eyes in wholemounted retinas. Optic nerves were used for either ETB western blot analysis (n = 24) or immunohistochemistry (n = 6) for ETB and glial fibrillary acidic protein (GFAP) to localize astrocytes. ETB expression was higher in the experimental nerve compared to the fellow untreated control nerve in 19 (79%) of the 24 animals with a mean increase of 16.7 +/- 4.5% in densitometric analyses of the immunoblots. Experimental nerves showed stronger labeling for both ETB and GFAP compared to control nerves. ETB-positive cells almost completely co-localized with GFAP-positive cells in both experimental and untreated control nerves, however, ETB expression was stronger in the astrocyte soma and proximal processes, while GFAP was expressed more strongly in the distal processes. There was a weak relationship between RGC loss and increase in ETB expression (r = -0.417, p = 0.076). There is an upregulation of ETB expression in optic nerve astrocytes in ET-1 induced chronic optic neuropathy causing RGC loss.


A precipitin for human serum proteins as released into the environment by stressed bait shrimp, Penaeus duorarum.
1. A precipitin for human serum proteins is released into the environment by stressed bait shrimp, Penaeus duorarum. 2. Two-dimensional crossed immunoelectrophoresis revealed that the precipitin reacts (a) primarily with proteins belonging to the major group, alpha-1 globulin; and (b) with more proteins than a standard mammalian antiserum. 3. This study extends the variety of species known to liberate precipitins and suggests this response may be widespread among invertebrates. 4. The precipitins are easily collected in saline solution and, by virtue of their unique specificities, are potentially useful in diagnostic testing.


Modulation of nitric oxide improves cyclosporin A-induced hypertension in rats and primates.
Cyclosporin-induced hypertension is a major complication of immunosuppression in transplant recipients but its pathophysiology is only partly understood. Cyclosporin reduces endothelium-dependent vasodilation and increases endothelin synthesis and release, which may contribute to this hypertension. We examined the effects of: (1) nitric oxide enhancement with L-arginine administration and antagonism with N-nitro-L-arginine; and (2) chronic endothelin receptor blockade with the non-peptide endothelin receptor antagonist, bosentan, in two animal models of cyclosporin-induced hypertension. Cyclosporin, administered daily to female Wistar rats (10 mg/kg per day for 30 days, s.c.) and to marmosets (30 mg/kg per day for 20 days, p.o.) significantly elevated tail cuff systolic blood pressure (BP). L-arginine (250 mg/kg, in saline, i.p.), N-nitro-L-arginine (25 mg/kg, in saline, i.p.), bosentan (100 mg/kg, in arabic gum, p.o.) or vehicle were given daily to the rats during the last week of cyclosporin treatment. Marmosets received L-arginine (300 mg/kg, in water, p.o.), bosentan (100 mg/kg/day in arabic gum, p.o.) or vehicle for the last 7 days of cyclosporin treatment. L-arginine, but not saline alone significantly lowered BP in the cyclosporin-hypertensive rats from 129 +/- 2 mm Hg to 122 +/- 3 mm Hg (P < 0.05), and cyclosporin-hypertensive marmosets from 156 +/- 2 mm Hg to 139 +/- 4 mm Hg (P < 0.01). NOLA significantly increased systolic BP in cyclosporin-treated (from 133 +/- 2 mm Hg at week 3 to 142 +/- 3 mm Hg, P < 0.05) and control rats (from 124.0 +/- 2 mm Hg to 134 +/- 2 mm Hg, P < 0.05) indicating that nitric oxide synthesis in cyclosporin-hypertensive rats could be further antagonised. Bosentan, but not arabic gum alone, also lowered BP in the cyclosporin-hypertensive rats from 134 +/- 1 mm Hg to 122 +/- 3 mm Hg (P < 0.01), and cyclosporin-hypertensive marmosets from 156 +/- 2 mm Hg to 139 +/- 4 mm Hg (P < 0.01). These results support the roles of both increased endothelin synthesis and decreased nitric oxide activity in the pathogenesis of cyclosporin A-induced hypertension.


Outcomes of secondary intraocular lens implantation in the Infant Aphakia Treatment Study.
To report outcomes of secondary intraocular lens (IOL) implantation in the Infant Aphakia Treatment Study (IATS). Multicenter clinical practice. Secondary analysis of patients enrolled in a randomized clinical trial. Details regarding all secondary IOL surgeries conducted in children enrolled in the IATS were compiled. Visual outcomes, refractive outcomes, and adverse events at the age of 10½ years were evaluated. Comparisons were made with eyes that remained aphakic and with eyes randomized to primary IOL placement. The study included 114 infants, 57 in the aphakic group and 57 in the primary IOL group; 55 of 57 patients randomized to aphakia with contact lens correction were seen for the 10½-year study visit; 24 (44%) of 55 eyes had secondary IOL surgery. Median age at IOL surgery was 5.4 years (range 1.7 to 10.3 years). Mean absolute prediction error was 1.00 ± 0.70 diopters (D). At age 10½ years, the median logarithm of the minimum angle of resolution visual acuity (VA) was 0.9 (range 0.2 to 1.7), similar to VA in the 31 eyes still aphakic (0.8, range 0.1 to 2.9); the number of eyes with stable or improved VA scores between the 4½-year and 10½-year study visits was also similar (78% secondary IOL eyes; 84% aphakic eyes). For eyes undergoing IOL implantation after the 4½-year study visit (n = 22), the mean refraction at age 10½ years was -3.20 ± 2.70 D (range -9.90 to 1.10 D), compared with -5.50 ± 6.60 D (n = 53, range -26.50 to 3.00 D) in eyes with primary IOL (P = .03). Delayed IOL implantation allows a more predictable refractive outcome at age 10½ years, although the range of refractive error is still large.


Prenatal diagnosis of a familial normal euchromatic variant of dup(15)(q11.2q11.2) in a pregnancy with a favorable outcome.
We present prenatal diagnosis of a familial normal euchromatic variant of dup(15)(q11.2q11.2) in a pregnancy with a favorable outcome. A 32-year-old woman underwent elective amniocentesis at 17 weeks of gestation because of anxiety. Amniocentesis revealed a karyotype of 46,XX,dup(15)(q11.2q11.2). Simultaneous array comparative genomic hybridization (aCGH) analysis on the DNA extracted from uncultured amniocytes revealed the result of arr (1-22, X) × 2 with no genomic imbalance. Cytogenetic analysis of the parental bloods showed that the mother had a karyotype of 46,XX,dup(15)(q11.2q11.2), and the father had a karyotype of 46,XY. Prenatal ultrasound findings were unremarkable. A healthy 2948 g female baby was delivered at 39 weeks of gestation without any phenotypic abnormality. Cytogenetic analysis of the cord blood revealed a karyotype of 46,XX,dup(15)(q11.2q11.2). Prenatal diagnosis of dup(15)(q11.2q11.2) should include a differential diagnosis of a 15q11.2 (BP1-BP2) microduplication encompassing TUBGCP5, CYFIP1, NIPA2 and NIPA1, and aCGH analysis is useful for the differential diagnosis under such a circumstance.


Epigenomic technologies for precision oncology.
Epigenetic patterns in a cell control the expression of genes and consequently determine the phenotype of a cell. Cancer cells possess altered epigenomes which include aberrant patterns of DNA methylation, histone tail modifications, nucleosome positioning and of the three-dimensional chromatin organization within a nucleus. These altered epigenetic patterns are potential useful biomarkers to detect cancer cells and to classify tumor types. In addition, the cancer epigenome dictates the response of a cancer cell to therapeutic intervention and, therefore its knowledge, will allow to predict response to different therapeutic approaches. Here we review the current state-of-the-art technologies that have been developed to decipher epigenetic patterns on the genomic level and discuss how these methods are potentially useful for precision oncology.


The political economics of cancer drug discovery and pricing.
Drug discoveries can, when used appropriately, save lives. Since 1970, cancer death rates among people aged under 65 have halved in countries such as the USA and the UK. Despite pharmaceutical market imperfections and fears about the prices of new treatments, further progress should be possible during the 2020s. Anticancer medicine outlays account for 0.1-0.2% of the gross domestic product (GDP) of developed countries. Total cancer service spending typically stands at ∼0.8% of GDP. The affordability of these sums is a political calculation. Improvements in the efficiency of drug development and global access to effective therapies are desirable. However, from a public interest perspective, these goals should not be pursued in ways that understate the value of better treatment outcomes and threaten the funding available for ongoing innovation.


Hyperbaric oxygen modulates antioxidant enzyme activity in rat skeletal muscles.
In skeletal muscle the activity of the enzymatic antioxidants superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) is regulated in response to generation of reactive oxygen species (ROS). Increased activity of these enzymes is observed after repeated bouts of aerobic exercise, a potent stimulus for intracellular ROS production. Hyperbaric oxygen (HBO) inhalation also stimulates intracellular ROS production although the effects of HBO on skeletal muscle SOD, GPx and CAT activity have not been studied. We tested the hypothesis that SOD, GPx and CAT activity is modulated in skeletal muscles in response to acute and repeated HBO administration. In adult male rats acute HBO inhalation (60 mm at 3 atmospheres absolute) reduced catalase activity by approximately 51% in slow-twitch soleus muscles. Additionally, repeated HBO inhalation (twice daily for 28 days) increased Mn2+-superoxide dismutase activity by approximately 241% in fast-twitch extensor digitorum longus muscles. We conclude that both acute and repeated HBO inhalation can alter enzymatic antioxidant activity in skeletal muscles.


Inferring muscular ground patterns in Bivalvia: Myogenesis in the scallop Nodipecten nodosus.
Myogenesis is currently investigated in a number of invertebrate taxa using combined techniques, including fluorescence labeling, confocal microscopy, and 3D imaging, in order to understand anatomical and functional issues and to contribute to evolutionary questions. Although developmental studies on the gross morphology of bivalves have been extensively pursued, organogenesis including muscle development has been scarcely investigated so far. The present study describes in detail myogenesis in the scallop Nodipecten nodosus (Linnaeus, 1758) during larval and postmetamorphic stages by means of light, electron, and confocal microscopy. The veliger muscle system consists of an anterior adductor muscle, as well as four branched pairs of striated velum retractors and two pairs of striated ventral larval retractors. The pediveliger stage exhibits a considerably elaborated musculature comprising the velum retractors, the future adult foot retractor, mantle (pallial) muscles, and the anterior and posterior adductors, both composed of smooth and striated portions. During metamorphosis, all larval retractors together with the anterior adductor degenerate, resulting in the adult monomyarian condition, whereby the posterior adductor retains both myofiber types. Three muscle groups, i.e., the posterior adductor, foot retractor, and pallial muscles, have their origin prior to metamorphosis and are subsequently remodeled. Our data suggest a dimyarian condition (i.e., the presence of an anterior and a posterior adductor in the adult) as the basal condition for pectinids. Comparative analysis of myogenesis across Bivalvia strongly argues for ontogenetic and evolutionary independence of larval retractors from the adult musculature, as well as a complex set of larval retractor muscles in the last common bivalve ancestor.


Prospectively randomized trial of adjuvant active-specific immunotherapy for human colorectal cancer.
Over the last four years a guinea pig model of active-specific immunotherapy (ASI) with a syngeneic tumor cell:bacillus calmette-Guerin (BCG) vaccine was translated into a prospectively randomized, controlled clinical trial in patients with colorectal cancer. Primary tumors from patients undergoing standard surgical resection were dissociated enzymatically and cryopreserved by techniques that maintain cell viability. Patients with transmural extension of tumor or nodal metastases were randomized into groups treated by resection alone (control) or resection plus ASI. With a mean follow-up of 28 months (range, 14-24), only 3 of 20 treatment patients had recurrences and none have died, whereas 9 of 20 control patients had recurrences and 4 died. These differences are statistically significant and are sufficiently encouraging to warrant expansion of these studies into other research centers.


Study of colonic IgG Fc binding site in cultured epithelial cells.
Previously, we identified a unique binding site for the Fc region of IgG in goblet cells of the human small intestine and colon. Understanding of the regulation and biological role of the binding site would be enhanced if it could be studied in cultured cells. Thus, we now have searched colonic carcinoma cell lines for presence of the site and further characterized it in such cells. HT29 and HT29-18N2 (a mucin-secreting subclone) cells were capable of binding IgG. The binding was associated with mucus-producing cells only in the cultures, was enhanced by culture of the cells in galactose-containing medium (which favors mucin production) as compared to glucose-containing medium, and was much greater for the HT29-18N2 cells than for the HT29 cells. LS 174T cells did not bind IgG despite the presence of numerous mucin-containing cells, and Colo 205 and LoVo cell cultures had neither mucin-containing cells nor IgG binding; thus, IgG binding and mucin production are not inseparably linked. By use of monoclonal antibodies to three different molecular-size components of the binding site, we found that a > 200-kDa component probably is necessary but not sufficient for IgG binding, whereas 78-kDa and 110- to 140-kDa components are not necessary.