Atmospheric vs. anaerobic processing of metabolome samples for the metabolite profiling of a strict anaerobic bacterium, Clostridium acetobutylicum.
Well-established metabolome sample preparation is a prerequisite for reliable metabolomic data. For metabolome sampling of a Gram-positive strict anaerobe, Clostridium acetobutylicum, fast filtration and metabolite extraction with acetonitrile/methanol/water (2:2:1, v/v) at -20°C under anaerobic conditions has been commonly used. This anaerobic metabolite processing method is laborious and time-consuming since it is conducted in an anaerobic chamber. Also, there have not been any systematic method evaluation and development of metabolome sample preparation for strict anaerobes and Gram-positive bacteria. In this study, metabolome sampling and extraction methods were rigorously evaluated and optimized for C. acetobutylicum by using gas chromatography/time-of-flight mass spectrometry-based metabolomics, in which a total of 116 metabolites were identified. When comparing the atmospheric (i.e., in air) and anaerobic (i.e., in an anaerobic chamber) processing of metabolome sample preparation, there was no significant difference in the quality and quantity of the metabolomic data. For metabolite extraction, pure methanol at -20°C was a better solvent than acetonitrile/methanol/water (2:2:1, v/v/v) at -20°C that is frequently used for C. acetobutylicum, and metabolite profiles were significantly different depending on extraction solvents. This is the first evaluation of metabolite sample preparation under aerobic processing conditions for an anaerobe. This method could be applied conveniently, efficiently, and reliably to metabolome analysis for strict anaerobes in air.


Postural Control While Walking Interferes With Spatial Learning in Older Adults Navigating in a Real Environment.
Cognitive demands for postural control increase with aging and cognitive-motor interference (CMI) exists for a number of walking situations, especially with visuo-spatial cognitive tasks. Such interference also influences spatial learning abilities among older adults; however, this is rarely considered in research on aging in spatial navigation. We posited that visually and physically exploring an unknown environment may be subject to CMI for older adults. We investigated potential indicators of postural control interfering with spatial learning. Given known associations between age-related alterations in gait and brain structure, we also examined potential neuroanatomical correlates of this interference. Fourteen young and 14 older adults had to find an invisible goal in an unfamiliar, real, ecological environment. We measured walking speed, trajectory efficiency (direct route over taken route) and goal fixations (proportion of visual fixations toward the goal area). We calculated the change in walking speed between the first and last trials and adaptation indices for all three variables to quantify their modulation across learning trials. All participants were screened with a battery of visuo-cognitive tests. Eighteen of our participants (10 young, 8 older) also underwent a magnetic resonance imaging (MRI) examination. Older adults reduced their walking speed considerably on the first, compared to the last trial. The adaptation index of walking speed correlated positively with those of trajectory efficiency and goal fixations, indicating a reduction in resource sharing between walking and encoding the environment. The change in walking speed correlated negatively with gray matter volume in superior parietal and occipital regions and the precuneus. We interpret older adults' change in walking speed as indicative of CMI, similar to dual task costs. This is supported by the correlations between the adaptation indices and between the change in walking speed and gray matter volume in brain regions that are important for navigation, given that they are involved in visual attention, sensory integration and encoding of space. These findings under ecological conditions in a natural spatial learning task question what constitutes dual tasking in older adults and they can lead future research to reconsider the actual cognitive burden of postural control in aging navigation research.


B Cell Receptor Repertoire Analysis in Malaria-Naive and Malaria-Experienced Individuals Reveals Unique Characteristics of Atypical Memory B Cells.
Malaria, caused by parasites of the Plasmodium genus, is responsible for significant morbidity and mortality globally. Chronic Plasmodium falciparum exposure affects the B cell compartment, leading to the accumulation of atypical memory B cells (atMBCs). IgM-positive (IgM+) and IgG+ atMBCs have not been compared in-depth in the context of malaria, nor is it known if atMBCs in malaria-experienced individuals are different from phenotypically similar B cells in individuals with no known history of Plasmodium exposure. To address these questions, we characterized the B cell receptor (BCR) repertoire of naive B cells (NBCs), IgM+ and IgG+ classical MBCs (cMBCs), and IgM+ and IgG+ atMBCs from 13 malaria-naive American adults and 7 malaria-experienced Ugandan adults. Our results demonstrate that P. falciparum exposure mainly drives changes in atMBCs. In comparison to malaria-naive adults, the BCR repertoire of Plasmodium-exposed adults showed increased levels of somatic hypermutation in the heavy chain V region in IgM+ and IgG+ atMBCs, shorter heavy chain complementarity-determining region 3 (HCDR3) in IgG+ atMBCs, and increased usage of IGHV3-73 in IgG+ cMBCs and both IgM+ and IgG+ atMBCs. Irrespective of Plasmodium exposure, IgM+ atMBCs closely resembled NBCs, while IgG+ atMBCs resembled IgG+ cMBCs. Physicochemical properties of the HCDR3 seemed to be intrinsic to cell type and independent of malaria experience. The resemblance between atMBCs from Plasmodium-exposed and naive adults suggests similar differentiation pathways regardless of chronic antigen exposure. Moreover, these data demonstrate that IgM+ and IgG+ atMBCs are distinct populations that should be considered separately in future analyses. IMPORTANCE Malaria, caused by Plasmodium parasites, still contributes to a high global burden of disease, mainly in children under 5 years of age. Chronic and recurrent Plasmodium infections affect the development of B cell memory against the parasite and promote the accumulation of atypical memory B cells (atMBCs), which have an unclear function in the immune response. Understanding where these cells originate from and whether they are beneficial in the immune response to Plasmodium will help inform vaccination development efforts. We found differences in B cell receptor (BCR) properties of atMBCs between malaria-naive and malaria-experienced adults that are suggestive of divergent selection processes, resulting in more somatic hypermutation and differential immunoglobulin heavy chain V (IGHV) gene usage. Despite these differences, atMBCs from malaria-naive and malaria-experienced adults also showed many similarities in BCR characteristics, such as physicochemical properties of the HCDR3 region, suggesting that atMBCs undergo similar differentiation pathways in response to different pathogens. Our study provides new insights into the effects of malaria experience on the B cell compartment and the relationships between atMBCs and other B cell populations.


Intrinsic fluorescence of a truncated Bordetella pertussis adenylate cyclase expressed in Escherichia coli.
A truncated, 432 residue long, Bordetella pertussis adenylate cyclase expressed in Escherichia coli was analyzed for intrinsic fluorescence properties. The two tryptophans (Trp69 and Trp242) of adenylate cyclase, each situated in close proximity to residues important for catalysis or binding of calmodulin (CaM), produced overlapping fluorescence emission bands upon excitation at 295 nm. CaM, alone or in association with low concentrations of urea, induced important modifications in the spectra of adenylate cyclase such as shifts of the maxima and change in the shape of the bands. From these changes and from the fluorescence spectrum of a modified form of adenylate cyclase, in which a valine residue was substituted for Trp242, it was deduced that, upon binding of CaM to the wild-type adenylate cyclase, only the environment of Trp242 was affected. The fluorescence maximum of this residue, which is more exposed to the solvent than Trp69 in the absence of CaM, is shifted by 13 nm to shorter wavelength upon interaction of protein with its activator. Trypsin cleaved adenylate cyclase into two fragments, one carrying the catalytic domain, and the second carrying the CaM-binding domain (Ladant et al., 1989). The isolated peptides conserved most of the environment around their single tryptophan residues, as in the intact adenylate cyclase, which suggests that the two domains of truncated B. pertussis adenylate cyclase also conserved most of their three-dimensional structure in the isolated forms.


Local opsonization by secreted macrophage complement components. Role of receptors for complement in uptake of zymosan.
We have examined the role of macrophage (M phi plasma membrane receptors for the cleaved third complement component (iC3b; CR3) and mannosyl, fucosyl terminated glycoproteins (MFR) in uptake of unopsonized zymosan. Monoclonal antibodies against CR3, M1/70 (Mac-1) and MO1, each inhibited approximately 50% of uptake of 125I-zymosan by murine and human M phi, respectively. Yeast mannan inhibited 0-50% of zymosan uptake in various M phi, in parallel with their expression of MFR. We demonstrated that M phi were the source of C3 in our assay and that the activity of other components of the complement system, namely a C3 convertase, factor I, and a factor I cofactor were also present in serum-free cultures of human monocytes. Macrophage C3 was deposited rapidly, within 10 min, on the zymosan particles and mediated binding, ingestion, and stimulation of superoxide release in BCG-activated and thioglycollate-elicited peritoneal M phi via CR3. Local secretion of complement proteins by M phi themselves can therefore opsonize pathogens and cells able to activate the alternative pathway and effect their destruction.


Conservative extraction of over-represented extensible motifs.
The discovery of motifs in biosequences is frequently torn between the rigidity of the model on the one hand and the abundance of candidates on the other. In particular, the variety of motifs described by strings that include 'don't care' (dot) patterns escalates exponentially with the length of the motif, and this gets only worse if a dot is allowed to stretch up to some prescribed maximum length. This circumstance tends to generate daunting computational burdens, and often gives rise to tables that are impossible to visualize and digest. This is unfortunate, as it seems to preclude precisely those massive analyses that have become conceivable with the increasing availability of massive genomic and protein data. Although a part of the problem is endemic, another part of it seems rooted in the various characterizations offered for the notion of a motif, that are typically based either on syntax or on statistics alone. It seems worthwhile to consider alternatives that result from a prudent combination of these two aspects in the model. We introduce and study a notion of extensible motif in a sequence which tightly combines the structure of the motif pattern, as described by its syntactic specification, with the statistical measure of its occurrence count. We show that a combination of appropriate saturation conditions (expressed in terms of minimum number of dots compatible with a given list of occurrences) and the monotonicity of probabilistic scores over regions of constant frequency afford us significant parsimony in the generation and testing of candidate over-represented motifs. The merits of the method are documented by the results obtained in implementation, which specifically targeted protein sequence families. In all cases tested, the motif reported in PROSITE as the most important in terms of functional/structural relevance emerges among the top 30 extensible motifs returned by our algorithm, often right at the top. Of equal importance seems the fact that the sets of all surprising motifs returned in each experiment are extracted faster and come in much more manageable sizes than would be obtained in the absence of saturation constrains. This software will be available for use with the suite of tools at www.research.ibm.com/bioinformatics.


Pharmacological characterization of [trans-5'-(4-amino-7,7-dimethyl-2-trifluoromethyl-7H-pyrimido[4,5-b][1,4]oxazin-6-yl)-2',3'-dihydrospiro(cyclohexane-1,1'-inden)-4-yl]acetic acid monobenzenesulfonate (JTT-553), a novel acyl CoA:diacylglycerol transferase (DGAT) 1 inhibitor.
Acyl CoA:diacylglycerol acyltransferase (DGAT) 1 is an enzyme that catalyzes the final step in triglyceride (TG) synthesis. This enzyme is considered to be a potential therapeutic target for obesity and diabetes. Here, results of an investigation of the pharmacological effects of JTT-553 [trans-5'-(4-amino-7,7-dimethyl-2-trifluoromethyl-7H-pyrimido[4,5-b][1,4]oxazin-6-yl)-2',3'-dihydrospiro(cyclohexane-1,1'-inden)-4-yl]acetic acid monobenzenesulfonate, a novel DGAT1 inhibitor, are reported. To measure the inhibitory activity of JTT-553 against DGAT1, TG synthesis using [(14)C]-labeled oleoyl-CoA was evaluated. Similarly, the inhibitory activity of JTT-553 against DGAT2, an isozyme of DGAT1, and acyl-CoA cholesterol acyltransferase (ACAT) 1, which is highly homologous to DGAT1, were evaluated. JTT-553 selectively inhibited human DGAT1 and showed comparable inhibitory effects on the activity of human, rat, and mouse DGAT. In vivo, JTT-553 suppressed plasma TG and chylomicron TG levels after olive oil loading in Sprague-Dawley (SD) rats. JTT-553 also inhibited TG synthesis in epididymal fat after [(14)C] oleic acid injection in C57BL/6J mice. Food intake was evaluated in SD rats fed 3.1%, 13%, or 35% (w/w) fat diets. In rats fed the 35% fat diet, JTT-553 reduced food intake. This reduction of food intake was observed 2 h after feeding, lasted for 24 h, and correlated with dietary fat content. Furthermore, JTT-553 reduced daily food intake and body weight gain in diet-induced obese rats after 4-week repeated administration. JTT-553 exerted multiple effects on intestinal fat absorption, adipose fat synthesis, and food intake, and consequently induced body weight reduction. Therefore, JTT-553 is expected to be an effective novel therapeutic agent for the treatment of obesity.


[Besides fibrinogen, are haemostatic, coagulation and/or fibrinolytic parameters predictors or markers of the risk of cardiovascular events?].
Most cardiovascular events result from a thrombotic complication of atherosclerotic lesions. In arterial vessels such as the coronary bed, an interrelationship of haemostatic, coagulation and fibrinolytic factors is implicated. While it can be demonstrated that fibrinogen is a risk factor/marker, the role of other factors is not well established. Under arterial flow conditions, platelets are predominantly involved in the thrombotic reaction. Yet, apart from a large increase in the platelet count, the involvement of platelet parameters in cardiovascular risk is not clearly evident. The lack of definitive platelet markers is at least partly due to the difficulty of studying platelet function ex vivo. Several polymorphisms of platelet glycoproteins carrying a moderate increase in risk have been reported, but only in younger patients. One potentially important factor for coagulation is the fibrin structure, which is dependent on fibrinogen, the rate of thrombin generation, the activity of factor XIII and the interrelationship of the cells concerned, all of which act on its sensitivity to thrombosis. Coagulation factors largely affect the rate of thrombin generation. The activity of the fibrinolytic system (and principally any deficiency) has a role in the cardiovascular risk. General markers of cardiovascular risk such as D-dimers are potentially useful, but they increase with thrombin generation and are decreased by a deficiency in fibrinolysis. Furthermore, possibly because they are not indicative of the fibrin structure, they are poorly correlated with clinical events. The poor significance of the available haemostatic, coagulation and/or fibrinolytic parameters is probably due to their lack of representativeness, since haemostatic, coagulation and fibrinolytic systems are all involved in the thrombotic response (and some in atherogenesis itself). Atherogenesis is a multifactorial process and numerous moderate risk factors act in association. Better predictability of the haemostatic tests would probably result from both the design of more representative tests and the evaluation of multiple parameters, and would lead to the definition of a risk score. Technological advances will make this possible.


Seedling mortality during biphasic drought in sandy Mediterranean soils.
Climate change is increasing the frequency and intensity of drought, and seedling response to a recurrent pattern of drought stress is necessary to understand vegetation establishment patterns in particularly for ecological restoration and conservation projects. A controlled environment study investigated seedling physiological response of framework Mediterranean tree species to simulated successive droughts. Six-month-old seedlings were grown in 1.0m tall pots to emulate deep soil profiles and subjected to a well watered treatment and a drought treatment consisting of an initial 60 day drought (water withholding), followed by 120 days of re-watering and a subsequent 60 day drought. Soil water access, soil water content, maximum root depth and xylem water potential were assessed through successive harvests. To assess seedling response to multiple droughts, gas-exchange and chlorophyll fluorescence measurements were taken every 15 days after each drought, and multiple times throughout re-watering. No seedling mortality was observed during the initial drought, whereas 100% mortality of all species occurred within 48 days of the second drought. Seedling gas exchange and water potential decreased with decreasing water availability but was dependent on the isohydric or anisohydric behaviour of individual species. An absence of sustained photoprotection during the second drought phase heightened photodamage to foliar tissues resulting in a more rapid decrease of gs and leaf water potential. Therefore, biphasic drought proved detrimental to seedling establishment by reducing physiological resilience, highlighting the severity of future climate change predictions towards the regeneration capacity of Mediterranean ecosystems.


Polymorphisms of interleukin-10 and its receptor and lung function in COPD.
Interleukin (IL)-10 is a type-2 T-helper cell cytokine with a broad spectrum of anti-inflammatory actions. Inflammation plays an important role in the pathogenesis of chronic obstructive pulmonary disease. It was hypothesised that single nucleotide polymorphisms (SNPs) of the genes encoding IL-10 (IL10) and the alpha subunit of its receptor (IL10RA) are associated with changes in, or value of, forced expiratory volume in one second (FEV1) in smoking-induced chronic obstructive pulmonary disease. In total, eleven SNPs of IL10 and IL10RA were studied in 586 White subjects, selected from continuous smokers followed for 5 yrs in the Lung Health Study, who showed the fastest (n=280) and slowest (n=306) decline in FEV1. These 11 SNPs were also studied in 1,072 participants exhibiting the lowest (n=538) and highest (n=534) baseline FEV1 at the beginning of the Lung Health Study. No association was found in the primary analyses. Although a subgroup analysis showed that the IL-10 3368A allele was associated with a fast decline in FEV1, the association did not pass correction for multiple comparisons. No gene-gene interaction of IL10 with IL10RA was found. There was no association of polymorphisms of the genes encoding interleukin-10 and the alpha subunit of its receptor with the rate of decline in, or value of, forced expiratory volume in one second in smoking-induced chronic obstructive pulmonary disease.


p27(Kip1)is overexpressed in very early stages of hepatocarcinogenesis.
Hepatocellular carcinoma (HCC) associated with chronic liver disease evolves from precancerous lesions and early HCC to more malignant forms. Despite the demonstrated importance of cell-cycle regulators in tumor biology, there have been few studies of their role in multistep hepatocarcinogenesis. Expression of p27(Kip1) and a degradation pathway associated protein, S-phase kinase-interacting protein 2 (Skp2), was therefore evaluated in surgically resected specimens of eight adenomatous hyperplasias, 16 early HCC and 126 classical HCC. Immunohistochemistry revealed no p27(Kip1) expression in the majority of hepatocytes from normal and cirrhotic liver, whereas positive staining for p27(Kip1) protein was found in 75.0% and 93.8% of adenomatous hyperplasias and early HCC, respectively. The average p27(Kip1) labeling indices (LI) for adenomatous hyperplasias, early HCC, well differentiated HCC, moderately differentiated HCC and poorly differentiated HCC were 36.99, 43.59, 47.73, 49.24, and 30.21, respectively. Real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) analyses confirmed the increases. Skp2 LI were also significantly elevated in accordance with stepwise progression of hepatocarcinogenesis. Increased expression of Skp2 mRNA was observed most frequently in less differentiated tumors and Kaplan-Meier survival analysis showed a significantly association with a poor prognosis (P = 0.0496). In conclusion, a high level of p27(Kip1) expression is evident from early stages of hepatocarcinogenesis, indicating that this parameter could be a useful diagnostic marker for precancerous lesions and early HCC. In addition, Skp2 expression correlates with tumor dedifferentiation and may contribute to biological aggression in HCC.


Leukemia inhibitory factor influences the timing of programmed synapses withdrawal from neonatal muscles.
We show that leukemia inhibitory factor (LIF) plays a physiological role in the programmed withdrawal of synapses from neonatal muscles. First, LIF mRNA is present in embryonic skeletal muscle and is developmentally regulated. We detect high levels of LIF mRNA at embryonic day 17 (E17) in mouse hind leg muscles. The content of LIF mRNA falls 10-fold between E17 and birth and then remains low in the neonate and adult. The decrease in LIF mRNA in skeletal muscle coincides with the end of secondary myogenesis and the completion of the adult number of myofibers. Second, treatment of the mouse tensor fascia latae (TFL), a superficial muscle of the hind leg, with LIF from birth (100 ng/day), transiently delays the withdrawal of excess inputs from polyneuronally innervated myofibers by approximately 3 days. The midpoint of the process is shifted from 7.5 +/- 10.2 +/- 0.6 days of age. LIF treatment delays synapse withdrawal by altering its timing without an appreciable effect on its rate. Third, in mice homozygous for a disruption of the LIF gene, the midpoint in the reduction of multiply innervated TFL myofibers occurs 1 day earlier, at 6.5 +/- 0.5 days of age. Muscle fiber number is unchanged in LIF null mice. Treatment with LIF does not alter the rate of neonatal growth, the number of muscle fibers in the TFL, or the reappearance of inputs that have been eliminated. Instead, LIF appears to delay maturation of the motor unit by transiently delaying the onset of synapse withdrawal. We hypothesize that this is a necessary component of a selective process that will operate simultaneously and equally on multiple, competing motor units.


[Social and hygienic factors influencing health state of doctors].
The article represents results of analysis concerning social and economic factors influencing health state of doctors in Bashkortostan Republic. The data were obtained through questioning of 6077 doctors. Findings are that over 75.0% of the doctors in the Republic live on verge of subsistence wage, 22% of the doctors have inadequate housing conditions; the doctors demonstrate low level of social optimism due to unsatisfactory economic conditions, social insecurity and high anxiety about the children's future.


Alzheimer's disease under managed care: implications from Medicare utilization and expenditure patterns.
Little information is available about the costs, utilization patterns, and the delivery system used by Medicare beneficiaries with chronic illnesses. This information will become increasingly important as more Medicare beneficiaries with chronic illness enroll in managed care plans and delivery systems must be developed to meet their needs. To analyze health care expenditures and utilization patterns for Medicare beneficiaries with dementia of the Alzheimer type (DAT) and compare them with those of all Medicare beneficiaries. A cross-sectional study. Practices providing services to Medicare beneficiaries in the U.S. Aged Medicare beneficiaries with DAT in fiscal year (FY) 1992. Medical expenditures and utilization patterns. In FY 1992, per capita Medicare expenditures for 9323 patients with DAT were $6208, or 1.9 times the per capita expenditure for all 1,221,615 beneficiaries in our sample. Inpatient care accounted for 62.7% of expenditures. Internal medicine was the specialty identified with the largest proportion of expenditures, but no single specialty accounted for the majority of care. Payments increased with comorbid conditions such as heart failure, chronic pulmonary diseases, and cerebrovascular disease. Current Medicare capitation payments to managed care plans may not meet the higher expected annual costs of care for beneficiaries with DAT. In turn, physicians (or physician groups) who accept capitation for Medicare beneficiaries with DAT should also consider how capitation rates are established by managed care plans and should learn ways to reduce financial risk.


Temperature-dependent development of the blow fly Chrysomya pinguis and its significance in estimating postmortem interval.
Chrysomya pinguis (Walker) (Diptera: Calliphoridae) is an endemic Asiatic blow fly species of forensic importance. Chrysomya pinguis is one of the first species to colonize a corpse, especially in high altitude areas during spring and autumn when the ambient temperature is lower. Despite its potential for forensic investigations to estimate the minimum postmortem interval (PMImin), little is known about the development of C. pinguis. In this study, C. pinguis was collected from the Yangtze River Delta region of China and reared at seven constant temperatures between 16°C and 34°C to investigate the effect of temperature on development duration, accumulated degree hours and larval body length of C. pinguis. Isomorphen and isomegalen diagrams for C. pinguis were generated using the results, and equations describing the variation in larval body length during development and the temperature-induced variation in development time were also obtained. Chrysomya pinguis can complete its life cycle at 16-34°C. The mean (±s.d.) developmental durations of C. pinguis from egg to adult at 16°C, 19°C, 22°C, 25°C, 28°C, 31°C and 34°C were 811.0 ± 3.8, 544.8 ± 2.0, 379.8 ± 1.8, 306.7 ± 2.4, 250.0 ± 2.8, 203.2 ± 2.1 and 185.3 ± 1.6 h, respectively. The mean (±s.e.) developmental threshold temperature D0 and the thermal summation constant K of the whole developmental process of C. pinguis were estimated as 10.88 ± 0.21°C and 4256.50 ± 104.50 degree hours, respectively. This study provides fundamental development data for the use of C. pinguis to estimate PMImin.


Associations between Theileria orientalis Ikeda type infection and the growth rates and haematocrit of suckled beef calves in the North Island of New Zealand.
The principle aim of this study was to examine the association between infection with Theileria orientalis Ikeda type and growth rates of suckled beef calves on four beef farms. In addition, associations between calf sex, sampling time, and individual farm and T. orientalis Ikeda type infection intensity and haematocrit (HCT) were investigated. The study was a prospective longitudinal study in which 240 calves from four purposively selected beef farms in the North Island of New Zealand were blood sampled and weighed in late spring, mid-summer and early autumn. Two farms were from high-risk (A and B) and two from low-risk (C and D) tick areas. Blood samples were analysed to determine HCT, and the number of T. orientalis Ikeda type organisms/µL of blood (infection intensity) using a quantitative PCR assay. A calf was defined as infected if >415 organisms/µL were detected in a blood sample. Linear mixed models were used to examine associations between infection intensity, mean daily liveweight gain (MDG), HCT, calf sex and time of sampling on the four farms. On Farms A and B nearly all calves were infected at each sampling time, on Farm C <30% were infected at any sampling and on Farm D infection prevalence increased from 32 to 79% between late spring and early autumn. On Farms C and D, from mid-summer to early autumn, mean MDG was 0.127 (95% CI=0.072-0.183) kg/day less for infected than uninfected calves (p<0.001). On all farms MDG was negatively associated with infection intensity for mid-summer and early autumn sampling times (p=0.037). The relationship between time of sampling and infection intensity varied between farms (p<0.001), and between male and female calves (p=0.018). Females had a higher infection intensity than males at the mid-summer and early autumn samplings. The association between HCT and infection intensity varied with sampling time and farm (p=0.018). There was a strong negative association between infection intensity and HCT at the late spring sampling, but in mid-summer there was no association, and in early autumn only a weak association. This study has shown that beef farmers in the North Island of New Zealand should be concerned about the welfare effects and economic impacts of T. orientalis Ikeda type infection in suckled beef calves.


[Planimetric evaluation of kinetic perimetry with the Goldman perimeter (author's transl)].
Kinetic perimetry was performed on 12 normal subjects. The perimetric results obtained with a Goldmann perimeter by 12 different investigators were statistically analyzed. Planimetry of areas outlined by a given isopter was done in order to reduce the amount of data. This led to a reduction in standard deviations of almost 50 percent. The statistical significance of field changes depends on individual variation and frequency of testing. If standard deviations are calculated in dB, variation becomes independent of given isopters. However, standard deviation may be increased by a factor of almost 2 as a result of the influence, of the investigator. Therefore, a standardization of test procedures by automatization seems meaningful, although standard deviations will not necessarily be lower with computerized perimetric methods.


Supracolloidal chains of patchy micelles of diblock copolymers with in situ synthesized nanoparticles.
Supracolloidal chains of diblock copolymer micelles were functionalized with gold and silver nanoparticles (NPs). Both NPs were independently synthesized in situ in the core of spherical micelles which were then converted to patchy micelles. With these patchy micelles as colloidal monomers, supracolloidal chains were polymerized by combining the patches of neighboring micelles. Since all micelles contained NPs, NPs were incorporated in every repeat unit of chains. In addition, a single gold NP was synthesized in the micellar core in contrast to several silver NPs so that we differentiated the chains with Au NPs from those with Ag NPs by the number of NPs in the repeat unit as well as by plasmonic bands in UV-Vis spectra.


The agreement chart.
When assessing the concordance between two methods of measurement of ordinal categorical data, summary measures such as Cohen's (1960) kappa or Bangdiwala's (1985) B-statistic are used. However, a picture conveys more information than a single summary measure. We describe how to construct and interpret Bangdiwala's (1985) agreement chart and illustrate its use in visually assessing concordance in several example clinical applications. The agreement charts provide a visual impression that no summary statistic can convey, and summary statistics reduce the information to a single characteristic of the data. However, the visual impression is personal and subjective, and not usually reproducible from one reader to another. The agreement chart should be used to complement the summary kappa or B-statistics, not to replace them. The graphs can be very helpful to researchers as an early step to understand relationships in their data when assessing concordance.


DDX3 enhances oncogenic KRAS‑induced tumor invasion in colorectal cancer via the β‑catenin/ZEB1 axis.
DDX3 plays a dual role in colorectal cancer; however, the role and underlying mechanism of DDX3 in colorectal tumorigenesis remains unclear. Here, we provide evidence that DDX3 enhances oncogenic KRAS transcription via an increase in SP1 binding to its promoter. Accelerating oncogenic KRAS expression by DDX3 promotes the invasion capability via the ERK/PTEN/AKT/β-catenin cascade. Moreover, the β-catenin/ZEB1 axis is responsible for DDX3-induced cell invasiveness and xenograft lung tumor nodule formation. The xenograft lung tumor nodules induced by DDX3-overexpressing T84 stable clone were nearly suppressed by the inhibitor of AKT (perifosine) or β-catenin (XAV939). Among patients, high KRAS, positive nuclear β-catenin expression and high ZEB1 were more commonly occurred in high-DDX3 tumors than in low-DDX3 tumors. High-DDX3, high-KRAS, positive nuclear β-catenin tumors, and high-ZEB1 exhibited worse overall survival (OS) and relapse free survival (RFS) than their counterparts. In conclusion, DDX3 may play an oncogenic role to promote tumor growth and invasion in colon cancer cells via the β-catenin/ZEB1 axis due to increasing KRAS transcription. We therefore suggest that AKT or β-catenin may potentially act as a therapeutic target to improve tumor regression and outcomes in colorectal cancer patients who harbored high-DDX3 tumors.


Cost of Caregivers for Treating Hospitalized Diarrheal Patients in Bangladesh.
Diarrheal diseases are a global public health problem and one of the leading causes of mortality, morbidity and economic loss. The objective of the study is to estimate the economic cost of caregivers and cost distribution per diarrheal episodes in Bangladesh. This was a cross-sectional hospital-based study conducted in public hospitals in Bangladesh. A total of 801 diarrheal patients were randomly selected and interviewed during January to December 2015. Simple descriptive statistics including frequencies, percentage, mean with 95% CI and median are presented. The overall average cost of caregivers was BDT 2243 (US$ 28.58) while only BDT 259 (US$ 3.29) was spent as out of pocket payments. Caregivers mostly spent money (US$ 1.63) for food, lodging, utility bills, and other lump sum costs followed by the transportation costs (US$ 1.57). The caregivers spent more (US$ 44.45) when they accompanied the patients who were admitted in inpatients care and almost 3.6 times higher than for out-patients care (US$ 12.42). The study delivers an empirical evidence to the health-care programmers and policy makers about the economic cost of caregivers during diarrheal treatment care, which should be accounted for in designing future diarrheal prevention programme.


SNP array genomic analysis of matched pairs of brain and liver metastases in primary colorectal cancer.
Brain metastasis formation is a rare and late event in colorectal cancer (CRC) patients and associated with poor survival. In contrast to other metastatic sites, the knowledge on chromosomal aberrations in brain metastases is very limited. Therefore, we carried out single nucleotide polymorphism (SNP) array analyses on matched primary CRC and brain metastases of four patients as well as on liver metastases of three patients. Brain metastases showed more chromosomal aberrations than primary tumors or liver metastases. Commonly occurring aberrations were gain of 8q11.1-q24.3 (primary CRC), gain of 13q12.13-q12.3 (liver metastases), and gain of 20q11.1-q13.33 (brain metastases). Furthermore, we found one copy-neutral loss of heterozygosity (cn-LOH) region on chromosome 3 in primary CRC, three cn-LOH regions in liver metastases and 23 cn-LOH regions in brain metastases, comprising 26 previously undescribed sites. The more frequent occurrence of cn-LOHs and subsequently affected genes in brain metastases shed light on the pathophysiology of brain metastasis formation. Further pairwise genetic analyses between primary tumors and their metastases will help to define the role of affected genes in cn-LOH regions.


Effects of different irrigation solutions on root dentine microhardness, smear layer removal and erosion.
This study aimed to compare the effects of different irrigants on root dentine microhardness, erosion and smear layer removal. A total of 72 root dentine slices were divided into six groups, according to the final irrigants used: Group 1: 17% ethylenediamine tetra-acetic acid (EDTA) + 2.5% NaOCl, Group 2: 7% maleic acid (MA) + 2.5% sodium hypochloride (NaOCl), Group 3: 1.3% NaOCl + mixture of tetracycline, acid and detergent (MTAD), Group 4: Smear Clear + 2.5% NaOCl, Group 5: 5% NaOCl, Group 6: saline. Vickers microhardness values were measured before and after treatment. In total, 42 root-halves were prepared for scanning electron microscope to evaluate the amount of smear and erosion in the coronal, middle and apical thirds. Data were analysed using two-way anova, Duncan and two-proportion z-tests. Maleic acid showed the greatest reduction in dentine microhardness (P < 0.05), followed by EDTA and MTAD. EDTA, maleic acid, MTAD and Smear Clear removed smear layer efficiently in the coronal and middle thirds of root canal. However, in the apical region, maleic acid showed more efficient removal of the smear layer than the other irrigants (P < 0.05).


Interleukin-11 attenuates pulmonary inflammation and vasomotor dysfunction in endotoxin-induced lung injury.
Interleukin (IL)-11, like other members of the gp130 receptor class, possesses anti-inflammatory properties. We hypothesized that IL-11 pretreatment would attenuate endotoxin [lipopolysaccharide (LPS)]-induced lung inflammation and diminish injury to endothelium-dependent and -independent mechanisms of pulmonary vasorelaxation that require cGMP in Sprague-Dawley rats. LPS (20 mg/kg ip) increased lung tumor necrosis factor (TNF)-alpha compared with the saline control (0.7 +/- 0.15 ng/g lung wet wt for control vs. 3.5 +/- 0.09 ng/g lung wet wt for LPS; P < 0.05). IL-11 (200 mg/kg ip) injected 10 min before LPS administration attenuated the LPS-induced lung TNF-alpha levels (1.6 +/- 0.91 ng/g lung wet wt; P < 0.05 vs. LPS). IL-11 also diminished LPS-induced lung neutrophil sequestration as assessed by myeloperoxidase units (2.1 +/- 0.25 U/g lung wet wt for saline and 15.6 +/- 2.02 U/g lung wet wt for LPS vs. 7.07 +/- 1.65 U/g lung wet wt for LPS plus IL-11; P < 0.05). Similarly, TNF-alpha binding protein (175 mg/kg) attenuated LPS-induced myeloperoxidase activity (6.04 +/- 0.14 U/g lung wet wt; P < 0.05). Both IL-11 and TNF-alpha binding protein similarly attenuated LPS-induced endothelium-dependent vasomotor dysfunction with improved relaxation responses to 10(-7) and 10(-6) M acetylcholine and A-23187 in phenylephrine-preconstricted isolated pulmonary artery rings (P < 0.05 vs. LPS). Endothelium-independent relaxation responses to sodium nitroprusside were also improved after LPS at 10(-6) M (P < 0.05 vs. LPS). Moreover, IL-11 decreased endotoxin-induced mortality in CF1 mice from 90 to 50% (P </= 0.05 vs. LPS). Therefore, IL-11 prevents LPS-induced lung TNF-alpha production, neutrophil sequestration, and pulmonary vasomotor dysfunction. We conclude that IL-11 possesses anti-inflammatory activity that protects against LPS-induced lung injury and lethality.


First-principles calculation of electronic structure of V-doped anatase TiO2.
The energetic and electronic structures of V-doped anatase TiO(2) have been investigated systematically by the GGA+U approach, including replacement of Ti by V in the absence and presence of oxygen vacancies and the presence of an interstitial site. It was found that V should exist as a V(4+) ion in the replacement of Ti in the anatase lattice, the electron transitions of which to the conduction band from V 3d states are responsible for the experimentally observed visible light absorption. The influence of V dopant concentration on the electronic and magnetic properties is also discussed, such as the influence of the U value in systems containing oxygen vacancies and spin flip phenomena for interstitial V-doping.


Engineering of hollow mesoporous silica nanoparticles for remarkably enhanced tumor active targeting efficacy.
Hollow mesoporous silica nanoparticle (HMSN) has recently gained increasing interests due to their tremendous potential as an attractive nano-platform for cancer imaging and therapy. However, possibly due to the lack of efficient in vivo targeting strategy and well-developed surface engineering techniques, engineering of HMSN for in vivo active tumor targeting, quantitative tumor uptake assessment, multimodality imaging, biodistribution and enhanced drug delivery have not been achieved to date. Here, we report the in vivo tumor targeted positron emission tomography (PET)/near-infrared fluorescence (NIRF) dual-modality imaging and enhanced drug delivery of HMSN using a generally applicable surface engineering technique. Systematic in vitro and in vivo studies have been performed to investigate the stability, tumor targeting efficacy and specificity, biodistribution and drug delivery capability of well-functionalized HMSN nano-conjugates. The highest uptake of TRC105 (which binds to CD105 on tumor neovasculature) conjugated HMSN in the 4T1 murine breast cancer model was ~10%ID/g, 3 times higher than that of the non-targeted group, making surface engineered HMSN a highly attractive drug delivery nano-platform for future cancer theranostics.


Using observational data to estimate an upper bound on the reduction in cancer mortality due to periodic screening.
Because randomized cancer screening trials are very expensive, observational cancer screening studies can play an important role in the early phases of screening evaluation. Periodic screening evaluation (PSE) is a methodology for estimating the reduction in population cancer mortality from data on subjects who receive regularly scheduled screens. Although PSE does not require assumptions about natural history of cancer it requires other assumptions, particularly progressive detection - the assumption that once a cancer is detected by a screening test, it will always be detected by the screening test. We formulate a simple version of PSE and show that it leads to an upper bound on screening efficacy if the progressive detection assumption does not hold (and any effect of birth cohort is minimal) To determine if the upper bound is reasonable, for three randomized screening trials, we compared PSE estimates based only on screened subjects with PSE estimates based on all subjects. In the three randomized screening trials, PSE estimates based on screened subjects gave fairly close results to PSE estimates based on all subjects. PSE has promise for obtaining an upper bound on the reduction in population cancer mortality rates based on observational screening data. If the upper bound estimate is found to be small and any birth cohort effects are likely minimal, then a definitive randomized trial would not be warranted.


Fura-2 fluorescence is localized to mitochondria in endothelial cells.
The new, highly fluorescent, calcium-sensitive dye, fura-2, can be loaded nondisruptively into intact cells by means of its permeant ester and used to measure the free calcium ion concentration in individual cells. For fura-2 to signal cytosolic calcium, it must be distributed homogeneously and exclusively throughout the cytoplasmic space. However, microscopic examination of bovine aortic endothelial cells loaded with fura-2 by exposure to its permeant ester reveals fluorescence associated with discrete intracellular structures rather than the homogeneous distribution expected for a cytosolic stain. Simultaneous labeling of bovine aortic endothelial cells with fura-2 and rhodamine 123 (a mitochondrial fluorescent vital stain) identifies these structures as mitochondria. Subcellular dye localizations are not observed when the cells are loaded with other putative cytosolic stains that gain access to the cytosol by means of a membrane permeant ester. Both carboxyfluorescein and indo-1 (another member of the family of second generation calcium indicators) stain the cytoplasm diffusely. It is suggested that fura-2 fluorescence accumulates in certain cells in association with mitochondria. It is important to assess the intracellular distribution of fura-2 when this indicator is used to measure the free cytosolic calcium ion concentration.


The Role of Compensation Criteria to Minimize Face-Time Bias and Support Faculty Career Flexibility: An Approach to Enhance Career Satisfaction in Academic Pathology.
Work-life balance is important to recruitment and retention of the younger generation of medical faculty, but medical school flexibility policies have not been fully effective. We have reported that our school's policies are underutilized due to faculty concerns about looking uncommitted to career or team. Since policies include leaves and accommodations that reduce physical presence, faculty may fear "face-time bias," which negatively affects evaluation of those not "seen" at work. Face-time bias is reported to negatively affect salary and career progress. We explored face-time bias on a leadership level and described development of compensation criteria intended to mitigate face-time bias, raise visibility, and reward commitment and contribution to team/group goals. Leaders from 6 partner departments participated in standardized interviews and group meetings. Ten compensation plans were analyzed, and published literature was reviewed. Leaders did not perceive face-time issues but saw team pressure and perception of availability as performance motivators. Compensation plans were multifactor productivity based with many quantifiable criteria; few addressed team contributions. Using these findings, novel compensation criteria were developed based on a published model to mitigate face-time bias associated with team perceptions. Criteria for organizational citizenship to raise visibility and reward group outcomes were included. We conclude that team pressure and perception of availability have the potential to lead to bias and may contribute to underuse of flexibility policies. Recognizing organizational citizenship and cooperative effort via specific criteria in a compensation plan may enhance a culture of flexibility. These novel criteria have been effective in one pilot department.


Characteristics of Corn Stover Components Pyrolysis at Low Temperature Based on Detergent Fibers.
In order to study the effect of biomass components on the low temperature (30-400°C) pyrolysis, the thermogravimetric analysis (TGA) of corn stover (CS) and its three detergent fibers (extracted by Van Soest method) were studied, and the model compounds of cellulose, hemicellulose and lignin were also tested as comparison. The results shows the low temperature pyrolysis index (P) is significantly different (PADL < Plignin < PCS < PNDF-CS < Pxylan < PADF-CS < Pcellulose). Klason-lignin has stronger thermal stability and decomposes more difficult than alkali lignin. The original cross-linked structure and interaction of the three components inhibited volatiles releasing, especially significant below 300°C. The synergistic effect between cellulose and lignin promoted devolatilization and decreased the initial temperature of cellulose decomposition. At last, the low temperature pyrolysis kinetics parameters (apparent activation energy and pre-exponential factor) of CS and its detergent fibers were calculated via the Coats-Redfern methods. This study can provide a theoretical basis for optimization of process conditions and industrial application of low temperature pyrolysis for lignocellulosic biomass.


Adenoma surveillance and colorectal cancer incidence: a retrospective, multicentre, cohort study.
Removal of adenomas reduces colorectal cancer incidence and mortality; however, the benefit of surveillance colonoscopy on colorectal cancer risk remains unclear. We examined heterogeneity in colorectal cancer incidence in intermediate-risk patients and the effect of surveillance on colorectal cancer incidence. We did this retrospective, multicentre, cohort study using routine lower gastrointestinal endoscopy and pathology data from patients who, after baseline colonoscopy and polypectomy, were diagnosed with intermediate-risk adenomas mostly (>99%) between Jan 1, 1990, and Dec 31, 2010, at 17 hospitals in the UK. These patients are currently offered surveillance colonoscopy at intervals of 3 years. Patients were followed up through to Dec 31, 2014.We assessed the effect of surveillance on colorectal cancer incidence using Cox regression with adjustment for patient, procedural, and polyp characteristics. We defined lower-risk and higher-risk subgroups on the basis of polyp and procedural characteristics identified as colorectal cancer risk factors. We estimated colorectal cancer incidence and standardised incidence ratios (SIRs) using as standard the general population of England in 2007. This trial is registered, number ISRCTN15213649. 253 798 patients who underwent colonic endoscopy were identified, of whom 11 944 with intermediate-risk adenomas were included in this analysis. After a median follow-up of 7·9 years (IQR 5·6-11·1), 210 colorectal cancers were diagnosed. 5019 (42%) patients did not attend surveillance and 6925 (58%) attended one or more surveillance visits. Compared to no surveillance, one or two surveillance visits were associated with a significant reduction in colorectal cancer incidence rate (adjusted hazard ratio 0·57, 95% CI 0·40-0·80 for one visit; 0·51, 0·31-0·84 for two visits). Without surveillance, colorectal cancer incidence in patients with a suboptimal quality colonoscopy, proximal polyps, or a high-grade or large adenoma (≥20 mm) at baseline (8865 [74%] patients) was significantly higher than in the general population (SIR 1·30, 95% CI 1·06-1·57). By contrast, in patients without these features, colorectal cancer incidence was lower than that of the general population (SIR 0·51, 95% CI 0·29-0·84). Colonoscopy surveillance benefits most patients with intermediate-risk adenomas. However, some patients are already at low risk after baseline colonoscopy and the value of surveillance for them is unclear. National Institute for Health Research Health Technology Assessment, Cancer Research UK.


Novel Schiff base cobalt(ii) phthalocyanine with appliance of MWCNTs on GCE: enhanced electrocatalytic activity behaviour of α-amino acids.
A novel tetra-4-{(E)-[(8-aminonaphthalen-1-yl)imino]methyl}-2-methoxyphenol Co(ii) phthalocyanine (CoTANImMMPPc) was synthesized using a precursor protocol and characterized via electroanalytical and spectroscopic techniques. The FT-IR spectra of the synthesized compounds showed significant peaks corresponding to the functional groups of the precursors and phthalocyanine (Pc) compound. The mass and NMR spectra confirmed the formation of the target precursor compounds. A film of CoTANImMMPPc was deposited on the surface of an electrode and applied for the detection and monitoring of l-alanine and l-arginine. The cyclic voltammetric studies of l-alanine and l-arginine using the (CoTANImMMPPc/MWCNTs/GC) electrode showed a linear response in the range of 50-500 nM and the limit of detection was found to be 1.5 and 1.2 nM, respectively. Differential pulse voltammetry and chronoamperometry showed that the catalytic response for l-alanine and l-arginine is in the range of 50-500 nM with an LoD of 1.8 and 2.3 nM, respectively. The oxidation-active CoTANImMMPPc film significantly enhanced the current response in the chronoamperometric method and displayed a selective and sensitive response towards l-alanine and l-arginine in the presence of various other bio-molecules. The developed electrode showed good working stability and was applied for the analysis of real samples, which yielded satisfactory results. Therefore, CoTANImMMPPc-MWCNTs/GCE shows good analytical performance, is economical and produced via a simple synthetic method and can be applied as a sensor for the detection of l-alanine and l-arginine.


Emerging structural insights into glycosyltransferase-mediated synthesis of glycans.
Glycans linked to proteins and lipids play key roles in biology; thus, accurate replication of cellular glycans is crucial for maintaining function following cell division. The fact that glycans are not copied from genomic templates suggests that fidelity is provided by the catalytic templates of glycosyltransferases that accurately add sugars to specific locations on growing oligosaccharides. To form new glycosidic bonds, glycosyltransferases bind acceptor substrates and orient a specific hydroxyl group, frequently one of many, for attack of the donor sugar anomeric carbon. Several recent crystal structures of glycosyltransferases with bound acceptor substrates reveal that these enzymes have common core structures that function as scaffolds upon which variable loops are inserted to confer substrate specificity and correctly orient the nucleophilic hydroxyl group. The varied approaches for acceptor binding site assembly suggest an ongoing evolution of these loop regions provides templates for assembly of the diverse glycan structures observed in biology.


Fresh Osteochondral Allograft Transplantation for Fractures of the Knee.
Objective The purpose of this study was to evaluate functional outcomes and allograft survivorship among patients with knee fracture who underwent fresh osteochondral allograft (OCA) transplantation as a salvage treatment option. Design Retrospective analysis of prospectively collected data. Setting Department of Orthopaedic Surgery at one hospital. Patients Fresh OCAs were implanted for osteochondral lesions after knee fracture in 24 males and 15 females with an average age of 34 years. Twenty-nine lesions (74%) were tibial plateau fractures, 6 (15%) were femoral condyle fractures, and 4 (10%) were patella fractures. Main Outcome Measurements Clinical evaluation included modified Merle d'Aubigné-Postel (18-point), International Knee Documentation Committee, and Knee Society function scores, and patient satisfaction. Failure of OCA was defined as revision OCA or conversion to total knee arthroplasty (TKA). Results Nineteen of 39 knees (49%) had further surgery. Ten knees (26%) were considered OCA failures (3 OCA revisions, 6 TKA, and 1 patellectomy). Survivorship of the OCA was 82.6% at 5 years and 69.6% at 10 years. Among the 29 knees (74%) that had the OCA still in situ, median follow-up was 6.6 years. Pain and function improved from preoperative to latest follow-up; 83% of patients reported satisfaction with OCA results. Conclusion OCA transplantation is a useful salvage treatment option for osteochondral lesions caused by knee fracture. Although the reoperation rate was high, successful outcome was associated with significant clinical improvement.


Timed adoptive T cell transfer during chemotherapy in patients with recurrent platinum-sensitive epithelial ovarian cancer.
The presence of T cells and suppressive myeloid cells in epithelial ovarian cancer (EOC) correlate with good and bad clinical outcome, respectively. This suggests that EOC may be sensitive to adoptive cell therapy with autologous tumor-infiltrating lymphocytes (TIL), provided that immunosuppression by myeloid-derived suppressor cells and M2 macrophages is reduced. Platinum-based chemotherapy can alleviate such immunosuppression, potentially creating a window of opportunity for T cell-based immunotherapy. We initiated a phase I/II trial (NCT04072263) in patients with recurrent platinum-sensitive EOC receiving TIL during platinum-based chemotherapy. TILs were administered 2 weeks after the second, third and fourth chemotherapy course. Patients were treated in two cohorts with or without interferon-α (IFNa), as conditioning and TIL support regimen. The primary endpoint was to evaluate the feasibility and safety according to CTCAE V.4.03 criteria and the clinical response and immune modulatory effects of this treatment were evaluated as secondary endpoints. Sixteen patients were enrolled. TIL could be successfully expanded for all patients. TIL treatment during chemotherapy without IFNa (n=13) was safe but the combination with IFNa added to the chemotherapy-induced toxicity with 2 out of 3 patients developing thrombocytopenia as dose-limiting toxicity. Fourteen patients completed treatment with a full TIL cycle and were further evaluated for clinical and immunological response. Platinum-based chemotherapy resulted in reduction of circulating myeloid cell numbers and IL-6 plasma levels, confirming its immunosuppression-alleviating effect. Three complete (CR), nine partial responses and two stable diseases were recorded, resulting in an objective response rate of 86% (Response Evaluation Criteria In Solid Tumors V.1.1). Interestingly, progression free survival that exceeded the previous platinum-free interval was detected in two patients, including an exceptionally long and ongoing CR in one patient that coincided with sustained alleviation of immune suppression. TIL therapy can be safely combined with platinum-based chemotherapy but not in combination with IFNa. The chemotherapy-mediated reduction in immunosuppression and the increase in platinum-free interval for two patients warrants further exploration of properly-timed TIL infusions during platinum-based chemotherapy, possibly further benefiting from IL-2 support, as a novel treatment option for EOC patients.


Post-Transplant Malignancy after Pediatric Kidney Transplantation: Retrospective Analysis of Incidence and Risk Factors in 884 Patients Receiving Transplants Between 1963 and 2015 at the University of Minnesota.
Post-transplant malignancy (PTM) remains a concern among pediatric kidney transplant (PKT) recipients. Between 1963 and 2015, 884 pediatric (age 0 to 17 years old) patients received 1,055 PKTs at our institution. Cox proportional hazards models were constructed to identify risk factors for PTM after PKT with time-to-first-PTM as a primary outcome. Secondly, the hazard of death or graft loss was calculated in patients who developed PTM. Median patient survival was 33 years (interquartile range [IQR] 18.7 to 47 years); 260 patients died during the study period and 47 had been diagnosed with PTM. There were 235 PTMs that occurred in 136 (15.4%) recipients at a median age of 29 years (IQR 17.8 to 37 years). The percentages of patients with PTM were 13% at 20 years post-PKT and 26% at 30 years post-PKT. Of PTM patients who died, 63.8% died of PTM. Among those who developed PTM, there was a higher hazard of death or graft loss (hazard ratio [HR] 1.62; 95% CI 1.11 to 2.38). In multivariable proportional hazards models, factors associated with PTM were increasing age at PKT (adjusted HR [AHR] 3.14; 95% CI 1.80 to 5.48 for 14 to 17 year-olds compared with children less than 3 years), having a living unrelated donor (LURD; AHR 3.25; 95% CI 1.27 to 8.35 compared with a living related donor), or implanting an Epstein-Barr virus (EBV)-positive allograft in an EBV-negative recipient (AHR 5.66; 95% CI 1.11 to 29.0). Compared with the general population, the cancer rate for PKT recipients was 6 times higher (126 vs 21 per 100,000 person-years). Pediatric kidney transplant recipients are at increased risk of PTM, which adversely affects survival. Children receiving transplants at an older age, from a LURD, or who receive an EBV-positive organ, should be monitored closely for the development of PTM.


PiMP my metabolome: an integrated, web-based tool for LC-MS metabolomics data.
The Polyomics integrated Metabolomics Pipeline (PiMP) fulfils an unmet need in metabolomics data analysis. PiMP offers automated and user-friendly analysis from mass spectrometry data acquisition to biological interpretation. Our key innovations are the Summary Page, which provides a simple overview of the experiment in the format of a scientific paper, containing the key findings of the experiment along with associated metadata; and the Metabolite Page, which provides a list of each metabolite accompanied by 'evidence cards', which provide a variety of criteria behind metabolite annotation including peak shapes, intensities in different sample groups and database information. PiMP is available at http://polyomics.mvls.gla.ac.uk, and access is freely available on request. 50 GB of space is allocated for data storage, with unrestricted number of samples and analyses per user. Source code is available at https://github.com/RonanDaly/pimp and licensed under the GPL. firstname.lastname@example.com. Supplementary data are available at Bioinformatics online.


Jump-Starting Faculty Development in Quality Improvement and Patient Safety Education: A Team-Based Approach.
Quality improvement (QI) and patient safety (PS) are cornerstones of health care delivery. Accreditation organizations increasingly require that learners engage in QIPS. For many faculty, these are new domains. Additional faculty development is needed for them to teach and mentor trainees. Existing programs, such as the Association of American Medical Colleges Teaching for Quality (Te4Q) program, target individual faculty and thus accommodate only limited participants at a time, which is problematic for institutions that need to train many faculty to support their learners. The authors invited diverse stakeholders from across the University of California, San Francisco (UCSF) School of Medicine and related health systems to participate in a team-based adaptation of the Te4Q program. The teams completed 5 projects based on previously identified priority areas to increase local capacity for QIPS teaching: (1) online modules for faculty new to QIPS, (2) a tool kit for graduate medical education programs, (3) a module for medical school clerkship directors, (4) guidelines for faculty to integrate early learners into QI projects, and (5) a "Teach-for-UCSF" certificate program in teaching QIPS. Thirty-five faculty members participated in the initial Te4Q workshop in January 2015, and by fall 2016, all projects were implemented. These projects led to additional faculty development initiatives and a rapidly expanding number of faculty across campus with expertise in teaching QIPS. Further collaborations between faculty focused on QIPS in care delivery and those focused on QIPS education to promote QIPS teaching have resulted from these initial projects.


IL-4 requirements for the generation of secondary in vivo IgE responses.
IL-4 has been shown to induce B lymphocytes to switch from the expression of membrane IgM to the expression of membrane IgE and to be required for the generation of primary polyclonal and secondary Ag-specific IgE responses in mice. To further define the role of IL-4 in the generation of memory IgE responses, we investigated the ability of a combination of anti-IL-4 and anti-IL-4R mAb to block the generation of secondary IgE responses induced by: 1) a second infection with the nematode parasites Nippostrongylus brasiliensis or Heligmosomoides polygyrus; or 2) injection of anti-IgD antibody-primed mice with anti-IgE antibody. The latter stimulus was designed to induce intrinsic membrane IgE-expressing B cells to differentiate into IgE-secreting cells. Although the IgE responses induced by a second nematode infection were completely inhibited by the combination of anti-IL-4 and anti-IL-4R mAb, anti-IgE antibody-induced IgE responses in anti-IgD primed mice were not inhibited by these antibodies to a large degree. Additional experiments demonstrated that the anti-IgE antibody-induced memory IgE response was dependent on CD4+ T cells but did not involve the low affinity B cell Fc epsilon RII. Taken together, these observations provide evidence that IL-4 is required for virgin B lymphocytes to develop into IgE-expressing cells, but is not required for B cells that express intrinsic membrane IgE to differentiate into IgE-secreting cells in a T-dependent response. Furthermore, these data suggest that secondary IgE responses in the parasite models that we have studied develop from B cells that had not previously switched to the expression of IgE.


Prognosis of white-coat and masked hypertension: International Database of HOme blood pressure in relation to Cardiovascular Outcome.
Home blood pressure monitoring is useful in detecting white-coat and masked hypertension and is recommended for patients with suspected or treated hypertension. The prognostic significance of white-coat and masked hypertension detected by home measurement was investigated in 6458 participants from 5 populations enrolled in the International Database of HOme blood pressure in relation to Cardiovascular Outcomes. During a median follow-up of 8.3 years, 714 fatal plus nonfatal cardiovascular events occurred. Among untreated subjects (n=5007), cardiovascular risk was higher in those with white-coat hypertension (adjusted hazard ratio 1.42; 95% CI [1.06-1.91]; P=0.02), masked hypertension (1.55; 95% CI [1.12-2.14]; P<0.01) and sustained hypertension (2.13; 95% CI [1.66-2.73]; P<0.0001) compared with normotensive subjects. Among treated patients (n=1451), the cardiovascular risk did not differ between those with high office and low home blood pressure (white-coat) and treated controlled subjects (low office and home blood pressure; 1.16; 95% CI [0.79-1.72]; P=0.45). However, treated subjects with masked hypertension (low office and high home blood pressure; 1.76; 95% CI [1.23-2.53]; P=0.002) and uncontrolled hypertension (high office and home blood pressure; 1.40; 95% CI [1.02-1.94]; P=0.04) had higher cardiovascular risk than treated controlled patients. In conclusion, white-coat hypertension assessed by home measurements is a cardiovascular risk factor in untreated but not in treated subjects probably because the latter receive effective treatment on the basis of their elevated office blood pressure. In contrast, masked uncontrolled hypertension is associated with increased cardiovascular risk in both untreated and treated patients, who are probably undertreated because of their low office blood pressure.


The British Pharmacological Society's WDM Paton Memorial Lecture 2019: How doctors were informed about pharmaceutical products through advertising in the British Medical Journal from 1955/6 to 1985/6.
We have reviewed pharmaceutical advertisements in every available issue of the British Medical Journal (BMJ) in 12-month periods during 1955/6, 1965/6, 1975/6, and 1985/6. We have determined the amount of advertising, the therapeutic areas covered, and whether adverts reflected the large number of New Chemical Entities (NCEs) launched during that time. For each product we recorded the therapeutic indications, the marketing company, and the number of adverts appearing. The total number of products advertised fell from 340 in 1955/6 to 260 in 1965/6, 70 in 1975/6, and 16 in 1985/6. Advertisement numbers and companies advertising also fell. Antimicrobial drugs and cardiovascular drugs were the top products advertised over the 30 years, with respiratory, analgesic, and gastrointestinal drugs also in the top five. The number of different drugs advertised by individual companies fell from around eight per company in 1955/6 to one or two in 1985/6. There was good concordance between the most advertised therapeutic areas and NCEs entering the market. From the 1950s to the 1980s prescribers were extensively informed about pharmacological advances in therapeutics through BMJ advertisements. Many novel drugs that were advertised proved to be of lasting value. The Medicines Act 1968 introduced product licensing, regulations requiring demonstration of quality, efficacy, and safety, and restrictions on advertising. Subsequently many companies reduced their advertising or stopped altogether. Since advertising influences prescribing, and since antimicrobial drugs were the most commonly advertised products during 1955-86, we speculate that advertising, resulting in excess use, may have, at least partly, driven bacterial drug resistance.


Chitinase-like proteins encoded in the genome of the pea aphid, Acyrthosiphon pisum.
In insects, chitinases play an essential role in the degradation of old exoskeleton and turnover of the gut lining. In silico screening of the entire genome of the pea aphid (Hemimetabola), Acyrthosiphon pisum, detected nine genes encoding putative chitinase-like proteins, including six enzymatically active chitinases, one imaginal disc growth factor, and one endo-beta-N-acetylglucosaminidase. Screening of the genomes of Aedes aegypti, Anopheles gambiae, Apis mellifera, Bombyx mori, Culex quinquefasciatus, Drosophila melanogaster, Nasonia vitripennis, Pediculus humanus corporis, and Tribolium castaneum suggested repeated gene duplications in holometabolous lineages. Quantitative reverse transcription-PCR demonstrated the expression of four and two distinct chitinase-like genes of A. pisum to be highly up-regulated in the embryo and the midgut, respectively, suggesting specific roles in these pea aphid tissues.


Polymorphisms in ABLIM1 are associated with personality traits and alcohol dependence.
Personality traits like novelty seeking (NS), harm avoidance (HA), and reward dependence (RD) are known to be moderately heritable (30-60%). These personality traits and their comorbidities, such as alcohol dependence (AD), may share genetic components. We examined 11,120 single nucleotide polymorphisms (SNPs) genotyped in 292 nuclear families from the Genetic Analysis Workshop 14, a subset from the Collaborative Study on the Genetics of Alcoholism (COGA). A family-based association analysis was performed using the FBAT program. NS, HA, and RD were treated as quantitative traits and AD as a binary trait. Based on a multivariate association test of three quantitative traits in FBAT, we observed 20 SNPs with p < 10(-3). Interestingly, several genes (TESK2, TIPARP, THEMIS, ABLIM1, RFX4, STON2 and LILRA1) are associated with three personality traits with p < 10(-3) using single trait analysis and AD. Especially, SNP rs727532 within ABLIM1 gene at 10q25 showed the most significant association (p = 6.4 × 10(-5)) in the multivariate test and strong associations with NS, HA, RD, and AD (p = 4.48 × 10(-4), 1.2 × 10(-5), 5.6 × 10(-5), 3.12 × 10(-4), respectively) in the COGA sample. In addition, the association of rs727532 with AD was confirmed in a replication study. This study reports some newly recognized associations between several genetic loci and both AD and three personality traits.


Asymmetry of gait after free flap reconstruction of severe tibial fractures with extensive soft-tissue damage.
Gait patterns of the preferred speed were investigated on 17 patients recovering (9 months-14 years) after reconstruction of severe tibial fractures. A novel data-acquisition system was used to record the plantar pressures as well as electromyographic (EMG) activities during walking. The results indicated incomplete recovery of symmetrical gait patterns. In particular, the duration of the stance phase was shorter on the operated side (mean (SD) 701 (90) ms compared with 765 (128) ms, p < 0.001). The peak pressure points under the foot were different on bilateral comparison, the operated side being regularly higher under the lateral forefoot area. This may imply attempts to reduce the loading of the ankle joint during stance. The pressure distribution models reflected these asymmetrical patterns more specifically than the EMG activities of the lower leg muscles examined.


Curriculum mapping of dental physiology curriculum: The path towards outcome-based education.
Curriculum mapping provides a clear picture of curriculum content, learning opportunities and assessment methods employed to measure the achievement of learning outcomes with their interrelationships. It facilitates educators and teachers to examine the extent to which the curricular components are linked and hence to find out gaps in the curriculum. The objective of the study was, therefore, to evaluate the physiology curriculum of Bachelor of Dental Surgery (BDS) programme through curriculum mapping. In this study, mapping of the physiology curriculum of three batches of BDS programme was conducted retrospectively. The components of the curriculum used for mapping were expected learning outcomes, curriculum content, learning opportunities, assessments and learning resources. The data were gathered by reviewing office records. Descriptive analysis of the data revealed reasonable alignment between the curriculum content and questions asked in examinations for all three batches. It was found that all the expected learning outcomes were addressed in the curriculum and assessed in different assessments. Moreover, the study revealed that the physiology curriculum was contributing to majority of the programme outcomes. Nevertheless, the study could identify some gaps in the curriculum, as well. This study revealed that majority of the components of the curriculum were linked and contributed to attaining the expected learning outcomes. It also showed that curriculum mapping was feasible and could be used as a tool to evaluate the curriculum.


Mutational analysis of the Tetrahymena telomerase RNA: identification of residues affecting telomerase activity in vitro.
Telomere-specific repeat sequences are essential for chromosome end stability. Telomerase maintains telomere length by adding sequences de novo onto chromosome ends. The template domain of the telomerase RNA component dictates synthesis of species-specific telomeric repeats and other regions of the RNA have been suggested to be important for enzyme structure and/or catalysis. Using enzyme reconstituted in vitro with RNAs containing deletions or substitutions we identified nucleotides in the RNA component that are important for telomerase activity. Although many changes to conserved features in the RNA secondary structure did not abolish enzyme activity, levels of activity were often greatly reduced, suggesting that regions other than the template play a role in telomerase function. The template boundary was only altered by changes in stem II that affected the conserved region upstream of the template, not by changes in other regions, such as stems I, III and IV, consistent with a role of the conserved region in defining the 5' boundary of the template. Surprisingly, telomerase RNAs with substitutions or deletion of residues potentially abolishing the conserved pseudoknot structure had wild-type levels of telomerase activity. This suggests that this base pairing interaction may not be required for telomerase activity per se but may be conserved as a regulatory site for the enzyme in vivo.


The Impact of Event Scale - Revised: psychometric properties of the Lithuanian version in a sample of employees exposed to workplace bullying.
The Impact of Event Scale-Revised (IES-R) is often used as a self-report instrument for symptoms of post-traumatic stress (PTS). The objective of this study was to explore the reliability and validity of the Lithuanian IES-R in a sample of employees with exposure to workplace bullying in different occupations. The original IES-R was translated into Lithuanian, and the comparability of content was verified through back-translation procedures. 294 employees with exposure to workplace bullying (52 teachers from the secondary schools of Kaunas, 56 family physicians, 101 nurses of internal medicine departments, 40 waiters, and 45 seafarers) were administered the Lithuanian IES-R and the General Health Questionnaire - 12 (GHQ-12) in order to verify some aspects of convergent validity. The exploratory factor analysis was used to verify the construct validity of the IES-R. The reliability of the Lithuanian version of the IES-R was verified. Cronbach's α of the total scale was 0.95. Exploratory factor analysis showed a clear factor structure with three independent dimensions: intrusion, avoidance and hyperarousal. Cronbach's α for subscales of intrusion, avoidance and hyperarousal were 0.89, 0.85, and 0.88, respectively. The convergent validity was supported by positive correlations between the subscales (intrusion, avoidance, hyperarousal) and the GHQ-12. The results suggest that the self-reported Lithuanian IES-R is a valid instrument for assessing the dimensions of post-traumatic stress, has good psychometric properties, and may be applied in prolonged trauma-exposed populations.


Beclin 1 mitigates motor and neuropathological deficits in genetic mouse models of Machado-Joseph disease.
Machado-Joseph disease or spinocerebellar ataxia type 3, the most common dominantly-inherited spinocerebellar ataxia, results from translation of the polyglutamine-expanded and aggregation prone ataxin 3 protein. Clinical manifestations include cerebellar ataxia and pyramidal signs and there is no therapy to delay disease progression. Beclin 1, an autophagy-related protein and essential gene for cell survival, is decreased in several neurodegenerative disorders. This study aimed at evaluating if lentiviral-mediated beclin 1 overexpression would rescue motor and neuropathological impairments when administered to pre- and post-symptomatic lentiviral-based and transgenic mouse models of Machado-Joseph disease. Beclin 1-mediated significant improvements in motor coordination, balance and gait with beclin 1-treated mice equilibrating longer periods in the Rotarod and presenting longer and narrower footprints. Furthermore, in agreement with the improvements observed in motor function beclin 1 overexpression prevented neuronal dysfunction and neurodegeneration, decreasing formation of polyglutamine-expanded aggregates, preserving Purkinje cell arborization and immunoreactivity for neuronal markers. These data show that overexpression of beclin 1 in the mouse cerebellum is able to rescue and hinder the progression of motor deficits when administered to pre- and post-symptomatic stages of the disease.


Uncovering per- and polyfluoroalkyl substances (PFAS) with nontargeted ion mobility spectrometry-mass spectrometry analyses.
Because of environmental and health concerns, legacy per- and polyfluoroalkyl substances (PFAS) have been voluntarily phased out, and thousands of emerging PFAS introduced as replacements. Traditional analytical methods target a limited number of mainly legacy PFAS; therefore, many species are not routinely assessed in the environment. Nontargeted approaches using high-resolution mass spectrometry methods have therefore been used to detect and characterize unknown PFAS. However, their ability to elucidate chemical structures relies on generation of informative fragments, and many low concentration species are not fragmented in typical data-dependent acquisition approaches. Here, a data-independent method leveraging ion mobility spectrometry (IMS) and size-dependent fragmentation was developed and applied to characterize aquatic passive samplers deployed near a North Carolina fluorochemical manufacturer. From the study, 11 PFAS structures for various per- and polyfluorinated ether sulfonic acids and multiheaded perfluorinated ether acids were elucidated in addition to 36 known PFAS. Eight of these species were previously unreported in environmental media, and three suspected species were validated.


[14C]2-deoxyglucose uptake in ground squirrel brain during hibernation.
Autoradiographic patterns of [14C]2-deoxyglucose uptake are described throughout the brains of hibernating and euthermic ground squirrels. Autoradiographs of the brains of hibernating animals are generally homogeneous in comparison to euthermic animals; hence, the relative 2-deoxyglucose uptake (R2DGU) of gray to white matter for the majority of the 85 neural structures examined decreases during hibernation. Two categories of structures are identified as potentially important in hibernation: (1) structures that have the highest R2DGU during hibernation (cochlear nucleus, paratrigeminal nucleus, and superior colliculus) and (2) structures that undergo the least reduction in R2DGU in the transition from euthermia to hibernation (suprachiasmatic nucleus and lateral septal nucleus). The percentage of reduction in R2DGU that a structure undergoes in the transition from euthermia to hibernation is proportional to the R2DGU of that structure during euthermia. The suprachiasmatic, paratrigeminal, and cochlear nuclei undergo less of a reduction than would be predicted from this relationship and may be particularly important during hibernation. Sensory nuclei that receive primary afferent projections are among the structures with the highest R2DGU during hibernation. These metabolically active structures may be responsible for the sensitivity of the hibernator to environmental stimuli.


Increasing physical exercise levels: age-specific benefits of planning.
This study examines the differential age effects on physical exercise of two planning interventions, action planning (when, where, how) and coping planning (anticipating barriers, mental simulation of success scenarios), and examines the mediating mechanisms of the interventions. The study assigned the participants, 205 cardiac rehabilitation patients, to one of the intervention groups (action-planning only or combined-planning group) or to a control group. Baseline measurement and follow-up took place 2 months apart. The interventions enhanced physical exercise independently of age. Pretreatment coping planning was higher in older (65-82 years) than in younger (38-54 years) or middle-aged (55-64 years) participants. At Time 2, older participants were the only ones without further increase in coping planning. Advancement in coping planning partially mediated the effect of the intervention. Coping planning facilitates improvement of physical exercise. Implications of age differences in planning are discussed.


Digestibility of amino acid in full-fat canola seeds, canola meal, and canola expellers fed to broiler chickens and pigs.
Canola products including full-fat canola seeds (FFCS), canola meal (CM), and canola expellers (CE) have been used in diets for both broiler chickens and pigs. However, their ability to utilize the AA in canola products might be different from each other. Therefore, this study was conducted to compare the apparent ileal digestibility (AID) and standardized ileal digestibility (SID) of CP and AA in broiler chickens and growing pigs fed FFCS, CM, and CE. Three diets were prepared to contain FFCS, CM, or CE as a sole source of nitrogen. A nitrogen-free diet was prepared. In Exp. 1, a total of 272 twenty-one-day-old male broiler chickens with an initial BW of 932 ± 80.6 g were assigned to diets in a randomized complete block design with BW as a blocking factor. After 5 d of feeding, birds were euthanized by sodium pentobarbital, and ileal digesta samples were collected from distal two-third of the ileum. In Exp. 2, 16 barrows were surgically fitted with T-cannulas at the distal ileum. After 8-d recovery period, pigs (initial BW = 18.9 ± 1.17 kg) were divided into 4 blocks based on BW and assigned to a quadruplicate 4 × 2 incomplete Latin Square design with 4 diets and 2 periods. Each period consisted of 5-d adaptation and 2-d ileal digesta collection periods. Data were analyzed as a 2 × 3 factorial treatment arrangement with effects of species (broiler chickens or pigs) and diets (FFCS, CM, or CE). There were interactions (P < 0.05) between species and experimental diets in the AID of all indispensable AA except for Lys. The AID of indispensable AA in FFCS for broiler chickens was greater (P < 0.05) than for pigs. Broiler chickens also had greater (P < 0.05) AID of Arg, His, Leu, Phe, and Val in CM compared with pigs; however, there were no differences in the AID of indispensable AA in CE between broiler chickens and pigs. The basal ileal endogenous losses of CP and AA, except Trp, in pigs were greater (P < 0.05) than in broiler chickens. There were also interactions (P < 0.05) between species and experimental diets in the SID of all indispensable AA except for Lys. Broiler chickens fed the diet containing FFCS had greater (P < 0.05) SID of indispensable AA compared with pigs fed the same diet; however, the SID of indispensable AA in CM or CE were not different between broiler chickens and pigs. In conclusion, differences in digestibility of AA in canola products were affected by nonruminant animal species.


LMO4 functions as a negative regulator of sensory organ formation in the mammalian cochlea.
In mammals, formation of the auditory sensory organ (the organ of Corti) is restricted to a specialized area of the cochlea. However, the molecular mechanisms limiting sensory formation to this discrete region in the ventral cochlear duct are not well understood, nor is it known whether other regions of the cochlea have the competence to form the organ of Corti. Here we identify LMO4, a LIM-domain-only nuclear protein, as a negative regulator of sensory organ formation in the cochlea. Inactivation of Lmo4 in mice leads to an ectopic organ of Corti (eOC) located in the lateral cochlea. The eOC retains the features of the native organ, including inner and outer hair cells, supporting cells, and other nonsensory specialized cell types. However, the eOC shows an orientation opposite to the native organ, such that the eOC appears as a mirror-image duplication to the native organ of Corti. These data demonstrate a novel sensory competent region in the lateral cochlear duct that is regulated by LMO4 and may be amenable to therapeutic manipulation.


Human amygdala reactivity is diminished by the β-noradrenergic antagonist propranolol.
Animal models of anxiety disorders emphasize the crucial role of locus ceruleus-noradrenergic (norepinephrine, NE) signaling, the basolateral amygdala (BLA) and their interactions in the expression of anxiety-like behavioral responses to stress. Despite clinical evidence for the efficacy of a β-noradrenergic receptor blockade with propranolol in the alleviation of anxiety symptoms and the secondary prevention of post traumatic stress disorder, preclinical evidence for a β-noradrenergic modulation of BLA activity in humans is missing. We combined functional magnetic resonance imaging in healthy volunteers with probabilistic mapping of intra-amygdalar responses to fearful, neutral and happy facial expressions to test the hypothesis that a β-noradrenergic receptor blockade with propranolol would inactivate the BLA. Consistent with our a priori hypothesis, propranolol diminished BLA responses to facial expressions, independent of their emotional valence. The absence of activity changes in probabilistically defined visual control regions underscores the specific action of propranolol in the BLA. Our findings provide the missing link between the anxiolytic potential of propranolol and the biological basis of β-noradrenergic activation in the human BLA as a key target for the pharmacological inhibition of anxiety neurocircuitry. Moreover, our findings add to emerging evidence that NE modulates both the reactivity (sensitivity) and the operating characteristics (specificity) of the BLA via β-noradrenergic receptors.


Roles of 5-hydroxytryptamine (5-HT) receptor subtypes in the inhibitory effects of 5-HT on C-fiber responses of spinal wide dynamic range neurons in rats.
5-Hydroxytryptamine (5-HT; serotonin) plays an important role in the descending control of nociception. 5-HT and its receptors have been extensively studied in the modulation of nociceptive transmission at the spinal level using behavioral tests that may be affected by the effects of 5-HT on motor performance and skin temperature. Using electrophysiological methods, the present study aimed to systematically investigate the roles of 5-HT receptor subtypes on the inhibitory effects of 5-HT on responses of the spinal wide dynamic range (WDR) neurons to C-fiber inputs in rats. Under basal conditions, topical application of 5-HT to the spinal cord inhibited the C-fiber responses of WDR neurons dose-dependently, whereas antagonists of 5-HT(1A) [WAY 100635 [N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinyl-cyclohexanecarboxamide maleate salt]], 5-HT(1B) [GR 55562 [3-[3-(dimethylamino)propyl]-4-hydroxy-N-[4-(4-pyrid-dinyl)phenyl]benzamide dihydrochloride]], 5-HT(2A) [ketanserin [3-[2-[4-(fluorobenzoyl)-1-piperidinyl]ethyl]-2,4[1H,3H]-quinazolinedione tartrate]], 5-HT(2C) [RS 102221 [8-[5-(2,4-dimethoxy-5-(4-trifluoromethylphenylsulfonamido)phenyl-5-oxopentyl]-1,3,8-triazaspiro[4.5]decane-2,4-dione hydrochloride]], 5-HT(3) [MDL 72222 [3-tropanyl-3,5-dichlorobenzoate]], and 5-HT(4) [GR 113808 ([1-[2-[(methylsulfonyl)-amino]ethyl]-4-piperidinyl]methyl 1-methyl-1H-indole-3-carboxylate)] had no effect on their own. The inhibitory effects of 5-HT were reversed by antagonists of 5-HT(1B) (GR 55562), 5-HT(2A) (ketanserin), 5-HT(2C) (RS 102221), 5-HT(3) (MDL 72222), and 5-HT(4) (GR 113808) but not by 5-HT(1A) (WAY 100635) receptor antagonists. Topical administration of agonists of 5-HT(1A) [(2R)-(+)-8-hydroxy-2-(di-n-propylamino)tetralin hydrobromide], 5-HT(1B) [CGS 12066 [7-trifluoromethyl-4-(4-methyl-1-piperazinyl)pyrrolo-[1,2-a]quinoxaline maleate salt]], 5-HT(2A) (alpha-methyl-5-hydroxytryptamine maleate), 5-HT(2C) [MK 212 [6-chloro-2-(1-piperazinyl)pyrazine hydrochloride]], 5-HT(3) [1-(3-chlorophenyl)biguanide hydrochloride], and 5-HT(4) [2-[1-(4-piperonyl)piperazinyl]benzothiazole] also inhibited the C-responses. These results suggest that, under basal conditions, there is no tonic serotonergic inhibition on the C-responses of dorsal horn neurons, and multiple 5-HT receptor subtypes including 1B, 2A, 2C, 3, and 4 may be involved in mediating the inhibitory effects of 5-HT.


[Carbamazepine hypersensitivity symptoms].
Carbamazepine (C) can cause a characteristic hypersensitivity reaction (CHS¿. This multisystem reaction typically presents as fever, mucocutaneous eruption and lymphadenopathy. The syndrome usually develops between 1 week and 3 months after starting therapy, with involvement of the liver, lung, kidney and inappropriate secretion of ADH. The incidence is less than 0.001% in those treated with C and it is diagnosed clinically. With onset of CHS, the drug must be stopped and if there is no improvement, cortico-steroids should be started. When the diagnosis is in doubt, the patch test, lymphocyte transformation test, macrophage migration inhibitor factor, and other tests can be helpful. The pathogenesis is not known. Similar syndromes have been described with phenytoin and phenobarbital. There is clinical and in-vitro evidence of cross reactions between C and phenytoin. It is not known whether the CHS syndrome should be considered a premalignant state, with increased risk for the development of malignant lymphoma.


TGF-beta1 release from biodegradable polymer microparticles: its effects on marrow stromal osteoblast function.
Controlled release of transforming growth factor-beta1 (TGF-beta1) to a bone defect may be beneficial for the induction of a bone regeneration cascade. The objectives of this work were to assess the feasibility of using biodegradable polymer microparticles as carriers for controlled TGF-beta1 delivery and the effects of released TGF-beta1 on the proliferation and differentiation of marrow stromal cells in vitro. Recombinant human TGF-beta1 was incorporated into microparticles of blends of poly(DL-lactic-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG). Fluorescein isothiocynate-labeled bovine serum albumin (FITC-BSA) was co-encapsulated as a porogen. The effects of PEG content (0, 1, or 5% by weight [wt%]) and buffer pH (3, 5, or 7.4) on the protein release kinetics and the degradation of PLGA were determined in vitro for as long as 28 days. Rat marrow stromal cells were seeded on a biodegradable poly(propylene fumarate) (PPF) substrate. The dose response and biological activity of released TGF-beta1 was determined after 3 days in culture. The effects of TGF-beta1 released from PLGA/PEG microparticles on marrow stromal cell proliferation and osteoblastic differentiation were assessed during a 21-day period. TGF-beta1 was encapsulated along with FITC-BSA into PLGA/PEG blend microparticles and released in a multiphasic fashion including an initial burst for as long as 28 days in vitro. Increasing the initial PEG content resulted in a decreased cumulative mass of released proteins. Aggregation of FITC-BSA occurred at lower buffer pH, which led to decreased release rates of both proteins. The degradation of PLGA was increased at higher PEG content and significantly accelerated at acidic pH conditions. Rat marrow stromal cells cultured on PPF substrates showed a dose response to TGF-beta1 released from the microparticles similar to that of added TGF-beta1, indicating that the activity of TGF-beta1 was retained during microparticle fabrication and after growth factor release. At an optimal TGF-beta1 dosage of 1.0 ng/ml after 3 days, the released TGF-beta1 enhanced the proliferation and osteoblastic differentiation of marrow stromal cells over 21 days of culture, with increased total cell number, alkaline phosphatase activity, and osteocalcin production. PLGA/PEG blend microparticles can serve as delivery vehicles for controlled release of TGF-beta1, and the released growth factor enhances marrow stromal cell proliferation and osteoblastic differentiation in vitro. Controlled release of TGF-beta1 from PLGA/PEG microparticles is representative of emerging tissue engineering technologies that may modulate cellular responses to encourage bone regeneration at a skeletal defect site.


Intracellular growth of Legionella pneumophila in Dictyostelium discoideum, a system for genetic analysis of host-pathogen interactions.
Conditions were established in which Legionella pneumophila, an intracellular bacterial pathogen, could replicate within the unicellular organism Dictyostelium discoideum. By several criteria, L. pneumophila grew by the same mechanism within D. discoideum as it does in amoebae and macrophages. Bacteria grew within membrane-bound vesicles associated with rough endoplasmic reticulum, and L. pneumophila dot/icm mutants, blocked for growth in macrophages and amoebae, also did not grow in D. discoideum. Internalized L. pneumophila avoided degradation by D. discoideum and showed evidence of reduced fusion with endocytic compartments. The ability of L. pneumophila to grow within D. discoideum depended on the growth state of the cells. D. discoideum grown as adherent monolayers was susceptible to L. pneumophila infection and to contact-dependent cytotoxicity during high-multiplicity infections, whereas D. discoideum grown in suspension was relatively resistant to cytotoxicity and did not support intracellular growth. Some known D. discoideum mutants were examined for their effect on growth of L. pneumophila. The coronin mutant and the myoA/B double myosin I mutant were more permissive than wild-type strains for intracellular growth. Growth of L. pneumophila in a G(beta) mutant was slightly reduced compared to the parent strain. This work demonstrates the usefulness of the L. pneumophila-D. discoideum system for genetic analysis of host-pathogen interactions.


Enalapril Influence on Arterial Stiffness in Rheumatoid Arthritis Women: A Randomized Clinical Trial.
Introduction: Cardiovascular parameters disruption can be found in patients at early stages of rheumatoid arthritis (RA). The primary endpoint of this study was the reduction of arterial stiffness in RA patients without traditional cardiovascular risk factors or previous comorbidities, measured by cardio-ankle vascular index (CAVI) through the enalapril intervention. The secondary endpoints were the enalapril influence on carotid femoral pulse wave velocity (cfPWV), carotid intima media thickness (cIMT), carotid artery distensibility (cDistensibility), Young's incremental elastic modulus (Einc)]. Materials and Methods: Fifty-three patients were enrolled in a clinical, randomized, closed-label trial. The subjects were randomly assigned into two groups: One receiving 5 mg of enalapril (27) or placebo (26), both twice a day. The drug was acquired at Victory Enterprises®. The placebo was kindly provided by the Universidad de Guadalajara (UdeG), as well as the blinding into two groups: A and B. Enalapril and placebo were packed into bottles without labeling. Clinical assessment included a structured questionnaire to gather demographic and clinical variables as well as determination of CAVI, cfPWV, cIMT, carotid artery distensibility and Einc. The whole set of evaluations were analyzed at the baseline and at the end of 12 weeks of intervention. Results: The CAVI measurement at baseline was 7.1 ± 1.4 and increased up to 7.5 ± 1.2 at the end of 12 weeks. Meanwhile, the enalapril group was as follows: 7.4 ± 1.2 and at the of intervention, reduced to 7.1 ± 0.9. A reduction in delta CAVI of 0.21 in the enalapril intervention group was found. In contrast, an increase of 0.39 was observed in the placebo group. The delta CAVI reduction was not influenced by age or peripheral systolic blood pressure (pSBP). Discussion: Enalapril seems to be effective in CAVI reduction in RA patients. The effect of enalapril intervention on arterial stiffness translated to the clinical context might be interpreted as a reduction of 6.4 years of arterial aging. Trial Registration: The protocol was approved by the Institutional Review Board with the register CI-0117 from UdeG, and 0211/18 from Hospital Civil "Dr. Juan I. Menchaca", Secretaría de Salud Jalisco: DGSP/DDI/D.INV.28/18 and retrospectively registered at ClinicalTrials.gov Protocol Registration and Results System: NCT03667131.