Wenshen Zhuanggu formula effectively suppresses breast cancer bone metastases in a mouse Xenograft model.
Wenshen Zhuanggu formula (WSZG) is a traditional Chinese medicine used as an adjuvant for the prevention of bone metastases in breast cancer patients. In this study we investigated the efficacy of WSZG in preventing bone metastases and the potential mechanisms in a mouse xenograft model of breast cancer bone metastases. This model was established by injection of human MDA-MB-231BO-Luc breast cancer cells alone or a mixture of the cancer cells with bone marrow-derived mesenchymal stem cells (BMSCs) into left ventricle of the heart in female nude mice. Then the mice were treated with WSZG (3.25, 6.5 or 13.0 mg·kg-1·d-1, ig) for four weeks, whereas zoledronic acid (100 μg/kg per week, ig) was used as a positive control. The occurrence and development of bone metastases were monitored via bioluminescent imaging, and bone lesions were assessed using micro-CT. Intracardiac injection of the mixture of MDA-MB-231BO-Luc breast cancer cells with BMSCs significantly facilitated the bone metastatic capacity of the breast cancer cells, and aggravated bone lesions in the mouse xenograft model of breast cancer bone metastases. Administration of WSZG dose-dependently inhibited the incidence and intensity of bone metastases and protected against bone lesions by suppressing osteoclast formation and tumor cell infiltration. Furthermore, administration of WSZG caused a marked reduction in the expression of CCL5/CCR5 and IL-17B/IL-17BR in bone metastatic tissues. The results demonstrate that WSZG exerts potential therapeutic effects in a mouse xenograft model of breast cancer bone metastases, which are partially mediated by weakening the interaction between BMSCs and breast cancer cells in the tumor microenvironment.


Social status, health status and therapy response in heroin addicts.
This study identified the impact of heroin compared to other drugs of abuse on the social, nutritional and health status of addicts, and assessed their response to detoxification. A total of 243 heroin addicts and 66 non-heroin addicts were included in the study. Heroin and non-heroin addiction had similar impact on social and nutritional status, but heroin addicts were seven times more likely to develop hepatitis B or C infection. Non-heroin addicts were more prone to develop depressed cellular immunity. The number of relapses was twice as high for heroin addicts who experienced more unpleasant symptoms during therapy. Active interventions to prevent and control addiction are needed regardless of the type of drug abused.


Thick trust, thin trust, social capital, and health outcomes among trans women of color in New York City.
Introduction: Many trans women of color communities experience high HIV seroprevalence, extreme poverty, high rates of victimization and substance use, and poor mental health. Greater knowledge of trans women of color social capital may contribute toward more effective services for this marginalized population. Methods: These data come from a mixed-methods study that examined trans/gender-variant people of color who attended transgender support groups at harm reduction programs in NYC. The study was conducted from 2011 to 12, total N = 34. The qualitative portion was derived from six focus group interviews. Results: Two support groups stood out as exhibiting very strong alternative kinship structures. One group was comprised of immigrant trans Latinas, and the other group were trans women of African descent living with HIV. Both groups demonstrated ample cultivation of "trust capital" in the form of "thick trust" (bonding capital) and "thin trust" (bridging/linking capital) both inside and outside/beyond the support groups. Thick trust included the cultivation of intimacy, support in primary romantic relationships, and community leadership. Thin trust included networking with a variety of organizations, increased educational opportunities, and cultural production. Discussion: Participants "opened up to social capital" through the process of trusting as a series of (1) risks; (2) vulnerabilities; and (3) reciprocities. A solid foundation of thick trust resulted in a social, psychological, and emotional "base." Upon this foundation, thin trust was operationalized resulting in positive material, economic, and quality-of-life outcomes, leading to an expanded space of capabilities.


Effects of sevelamer carbonate on advanced glycation end products and antioxidant/pro-oxidant status in patients with diabetic kidney disease.
The primary goals were to re-examine whether sevelamer carbonate (SC) reduces advanced glycation end products (AGEs) (methylglyoxal and carboxymethyllysine [CML]), increases antioxidant defenses, reduces pro-oxidants, and improves hemoglobin A1c (HbA1c) in patients with type 2 diabetes mellitus (T2DM) and diabetic kidney disease (DKD). Secondary goals examined albuminuria, age, race, sex, and metformin prescription. This two-center, randomized, intention-to-treat, open-label study evaluated 117 patients with T2DM (HbA1c >6.5%) and stages 2-4 DKD (urinary albumin/creatinine ratio ≥200 mg/g) treated with SC (1600 mg) or calcium carbonate (1200 mg), three times a day, without changing medications or diet. Statistical analyses used linear mixed models adjusted for randomization levels. Preselected subgroup analyses of sex, race, age, and metformin were conducted. SC lowered serum methylglyoxal (95% confidence interval [CI], -0.72 to -0.29; P<0.001), serum CML (95% CI, -5.08 to -1.35; P≤0.001), and intracellular CML (95% CI, -1.63 to -0.28; P=0.01). SC increased anti-inflammatory defenses, including nuclear factor like-2 (95% CI, 0.58 to 1.29; P=0.001), AGE receptor 1 (95% CI, 0.23 to 0.96; P=0.001), NAD-dependent deacetylase sirtuin-1 (95% CI, 0.20 to 0.86; P=0.002), and estrogen receptor α (95% CI, 1.38 to 2.73; P ≤0.001). SC also decreased proinflammatory factors such as TNF receptor 1 (95% CI, -1.56 to -0.72; P≤0.001) and the receptor for AGEs (95% CI, -0.58 to 1.53; P≤0.001). There were no differences in HbA1c, GFR, or albuminuria in the overall group. Subanalyses showed that SC lowered HbA1c in women (95% CI, -1.71 to -0.27; P=0.01, interaction P=0.002), and reduced albuminuria in those aged <65 years (95% CI, -1.15 to -0.07; P=0.03, interaction P=0.02) and non-Caucasians (95% CI, -1.11 to -0.22; P=0.003, interaction P≤0.001), whereas albuminuria increased after SC and calcium carbonate in Caucasians. SC reduced circulating and cellular AGEs, increased antioxidants, and decreased pro-oxidants, but did not change HbA1c or the albumin/creatinine ratio overall in patients with T2DM and DKD. Because subanalyses revealed that SC may reduce HbA1c and albuminuria in some patients with T2DM with DKD, further studies may be warranted.


Silver(I) double and multiple salts containing the 1,3-butadiynediide dianion: coordination diversity and assembly with the supramolecular synthon Ag4[cap]C[triple bond]C-C[triple bond]C[cap]Ag4.
A series of 13 silver(I) double and multiple salts containing 1,3-butadiynediide, C4(2-), were synthesized by dissolving the silver carbide Ag2C4 in a concentrated aqueous solution of one or more of the silver salts AgNO3, AgCF3CO2, AgC2F5CO2, AgF, AgBF4, and AgPF6. The 1,3-butadiynediide anion invariably adopts a mu4,mu4 coordination mode in these compounds, which indicates that the Ag4[cap]C[triple bond]C-C[triple bond]C[cap]Ag4 moiety can be used as a new type of metalloligand supramolecular synthon for the construction of coordination networks. Fine-tuning with various ancillary anionic ligands caused the Ag4 aggregate at each ethynide terminus to adopt a butterfly-shaped, planar, or barblike configuration, within which the silver-ethynide interactions can be classified into three types: sigma, pi, and mixed (sigma,pi). The effect of coexisting nitrile ligands and quaternary ammonium salts on supramolecular assembly with the above synthon was also explored. The hydrolysis of PF6(-) and BF4(-) led to the formation of the quadruple salt Ag2C4 x 4 AgNO3 x AgPF2O2 x Ag3PO4 and a novel (F)2(H2O)18 hydrogen-bonded tape in the triple salt Ag2C4 x 2 AgF x 10 AgC2F5CO2 x CH3CN x 12 H2O, respectively. The largest silver-ethynide cluster aggregate described to date, (C4)3@Ag18, occurs in 3 Ag2C4 x 12 AgC2F5CO2 x 5 [(BnMe3N)C2F5CO2] x 4 H2O (Bn = benzyl).


Activation of ADP-ribosyltransferase in polyamine-depleted mammalian cells.
Mammalian fibroblasts were cultured in the presence of alpha-methylornithine and/or methylglyoxal bis(guanylhydrazone), which inhibit the synthesis of polyamines. This led to a decrease in the cellular content of the polyamines spermine and spermidine by up to 60% when the cells were grown in the presence of both drugs together. The activity of the chromatin-associated enzyme ADP-ribosyltransferase was enhanced 2-3-fold in the drug-treated cells when measured in cells subsequently rendered permeable to exogenous NAD+, the substrate for the transferase. This is a novel and surprising observation, since the transferase is invariably activated by the addition of polyamines to a suitable incubation system such as permeabilized cells, isolated nuclei or the purified enzyme. We found no evidence that the activation was due to the appearance of DNA strand breaks, by using a variety of procedures including both neutral [the 'nucleoid' technique of Cook & Brazell [(1975) J. Cell Sci. 19, 261-279; (1976) J. Cell Sci. 22, 287-302]] and alkaline sucrose-gradient centrifugation and gel electrophoresis, suggesting that this therefore may not be the only means of regulating the activity of ADP-ribosyltransferase and that polyamines may have a role to play in this regard in vivo.


Levosimendan for patients with severely reduced left ventricular systolic function and/or low cardiac output syndrome undergoing cardiac surgery: a systematic review and meta-analysis.
Previous studies have shown beneficial effects of levosimendan in high-risk patients undergoing cardiac surgery. Two large randomized controlled trials (RCTs), however, showed no advantages of levosimendan. We performed a systematic review and meta-analysis (MEDLINE and Embase from inception until March 30, 2017), investigating whether levosimendan offers advantages compared with placebo in high-risk cardiac surgery patients, as defined by preoperative left ventricular ejection fraction (LVEF) ≤ 35% and/or low cardiac output syndrome (LCOS). The primary outcomes were mortality at longest follow-up and need for postoperative renal replacement therapy (RRT). Secondary postoperative outcomes investigated included myocardial injury, supraventricular arrhythmias, development of LCOS, acute kidney injury (AKI), duration of mechanical ventilation, intensive care unit and hospital lengths of stay, and incidence of hypotension during drug infusion. Six RCTs were included in the meta-analysis, five of which investigated only patients with LVEF ≤ 35% and one of which included predominantly patients with LCOS. Mortality was similar overall (OR 0.64 [0.37, 1.11], p = 0.11) but lower in the subgroup with LVEF < 35% (OR 0.51 [0.32, 0.82], p = 0.005). Need for RRT was reduced by levosimendan both overall (OR 0.63 [0.42, 0.94], p = 0.02) and in patients with LVEF < 35% (OR 0.55 [0.31, 0.97], p = 0.04). Among secondary outcomes, we found lower postoperative LCOS in patients with LVEF < 35% receiving levosimendan (OR 0.49 [0.27, 0.89], p = 0.02), lower overall AKI (OR 0.62 [0.42, 0.92], p = 0.02), and a trend toward lower mechanical support, both overall (p = 0.07) and in patients with LVEF < 35% (p = 0.05). Levosimendan reduces mortality in patients with preoperative severely reduced LVEF but does not affect overall mortality. Levosimendan reduces the need for RRT after high-risk cardiac surgery.


Comparing Outcomes of Critically Ill Patients in Intensive Care Units and General Wards: A Comprehensive Analysis.
The admission of critically ill patients to intensive care unit (ICU) plays a crucial role in reducing mortality. However, the scarcity of available ICU beds presents a significant challenge. In resource-limited settings, the outcomes of critically ill patients, particularly those who are not accepted for ICU admission, have been a topic of ongoing debate and contention. This study aimed to explore the outcomes and factors associated with ICU admission and mortality among critically ill patients in Thailand. This prospective cohort study enrolled critically ill adults indicated for medical ICU admission. Patients were followed for 28 days regardless of whether they were admitted to an ICU. Data on mortality, hospital length of stay, duration of organ support, and factors associated with mortality and ICU admission were collected. Of the 180 patients enrolled, 72 were admitted to ICUs, and 108 were cared for in general wards. The ICU group had a higher 28-day mortality rate (44.4% vs 20.4%; P=0.001), but other outcomes of interest were comparable. Multivariate analysis identified alteration of consciousness, norepinephrine use, and epinephrine use as independent predictors of 28-day mortality. Higher body mass index (BMI), higher APACHE II score, and acute kidney injury were predictive factors associated with ICU acceptance. Among patients indicated for ICU admission, those who were admitted had a higher 28-day mortality rate. Higher mortality was associated with alteration of consciousness and vasopressor use. Patients who were sicker and had higher BMI were more likely to be admitted to an ICU.


IAEA Activities on 67Cu, 186Re, 47Sc Theranostic Radionuclides and Radiopharmaceuticals.
Despite interesting properties, the use of 67Cu, 186Re and 47Sc theranostic radionuclides in preclinical studies and clinical trials is curtailed by their limited availability due to a lack of widely established production methods. An IAEA Coordinated Research Project (CRP) was initiated to identify important technical issues related to the production and quality control of these emerging radionuclides and related radiopharmaceuticals, based on the request from IAEA Member States. The international team worked on targetry, separation, quality control and radiopharmaceutical aspects of the radionuclides obtained from research reactors and cyclotrons leading to preparation of a standard recommendations for all Member States. The CRP was initiated in 2016 with fourteen participants from thirteen Member States from four continents. Extraordinary results on the production, quality control and preclinical evaluation of selected radionuclides were reported in this project that was finalized in 2020. The outcomes, outputs and results of this project achieved by participating Member States are described in this minireview.


Field testing of an enhanced direct-fecal polymerase chain reaction procedure, bacterial culture of feces, and a serum enzyme-linked immunosorbent assay for detecting Mycobacterium avium subsp paratuberculosis infection in adult dairy cattle.
To estimate the sensitivity (Se) and specificity (Sp) for an enhanced direct-fecal PCR procedure, bacterial culture of feces (BCF), and a serum ELISA for detecting Mycobacterium avium subsp paratuberculosis (MAP) infection in adult dairy cattle. Fecal and serum samples were collected from 669 adult cattle randomly selected from a 4,000-cow dairy herd known to contain animals infected with MAP. Serum samples were evaluated for MAP-specific antibodies via ELISA. Fecal samples were evaluated by BCF and enhanced PCR methods (both gel-based [GB]-PCR and quantitative real-time [qRT]-PCR assays). Fecal samples also were pooled (5:1) and then subjected to GB-PCR assay. Bayesian statistical methods were used to estimate Se and Sp for each diagnostic test without knowledge concerning true MAP infection status. Adjusting for Se conditional dependence between serum ELISA and BCR, overall Se and Sp were estimated at 33.7% and 95.9%, 51.3% and 99.0%, and 32.2% and 100% for serum ELISA, qRT-PCR, and BCF, respectively.The GB-PCR assay yielded positive results for 38.3% of the pools known to contain feces from at least 1 cow that had positive GBPCR results. Estimated Se values for the serum ELISA and BCF were slightly lower than those reported elsewhere. The enhanced qRT-PCR method offered relative improvements in Se of 52% and 59% over serum ELISA and microbial culture, respectively. Pooling of fecal samples and testing with the GB-PCR assay are not recommended. Additional studies with qRT-PCR and fecal pools are required.


Distribution of free and antibody-bound peptide hormones in two-phase aqueous polymer systems.
Peptide hormones labelled with radioactive iodine were partitioned into the aqueous two-phase polymer systems developed by Albertsson (1960) and the conditions required for separation of free from antibody-bound hormone have been worked out. Hormones studied included insulin, growth hormone, parathyroid hormone and [arginine]-vasopressin. Free and antibody-bound hormones show different distribution coefficients in a number of systems tested; two systems, the dextran-polyethylene glycol and dextran sulphate-polyethylene glycol system, give optimum separation. Free hormones distribute readily into the upper phase of these systems, whereas hormone-antibody complexes, as well as uncombined antibody, are found almost completely in the lower phase. Various factors including the polymer concentration, the ionic composition of the system, the nature of the hormone and the nature of added serum protein differentially affect the distribution coefficients for free and antibody-bound hormone. These factors can be adequately controlled so as to improve separation. The two-phase partition method has been successfully applied to measure binding of labelled hormone to antibody under standard radioimmunoassay conditions. It exhibits several advantages over the method of equilibration dialysis and can be applied to the study of non-immunological interactions.


Senile and postmenopausal osteoporosis.
Obviously, the relentless decrease in bone mass that accompanies aging will continue the long sought "elixir of youth" is discovered. Individuals, because of race, sex, environmental, dietary, genetic or activity differences, will be more or less predisposed to symptomatic osteoporosis with increasing age. The careful and knowledgeable physician should, however, make every attempt to rule out potentially remediable, subtle forms of demineralizing disorders, such as apathetic or T3-thyrotoxicosis, hyperparathyroidism, malabsorption and osteomalacia or multiple myeloma. Not only do these diseases result in an accelerated loss of bone mass and an increased incidence of skeletal fractures but they mimic postmenopausal or senile osteoporosis radiologically. Once the metabolic or malignant disorders of bone metabolism have been effectively considered and ruled out, the senescent or postmenopausal osteoporotic patient should be treated judiciously with short-term estrogen therapy, a diet sufficient in vitamin D and calcium content and continued attempts to insure adequate skeletal mobilization. The addition of sodium fluoride and/or calcitonin to this regimen should not be attempted without extreme caution until the potentially harmful systemic effects of prolonged therapeutic trials have been appropriately assessed.


Self-terminating diffraction gates femtosecond X-ray nanocrystallography measurements.
X-ray free-electron lasers have enabled new approaches to the structural determination of protein crystals that are too small or radiation-sensitive for conventional analysis1. For sufficiently short pulses, diffraction is collected before significant changes occur to the sample, and it has been predicted that pulses as short as 10 fs may be required to acquire atomic-resolution structural information1-4. Here, we describe a mechanism unique to ultrafast, ultra-intense X-ray experiments that allows structural information to be collected from crystalline samples using high radiation doses without the requirement for the pulse to terminate before the onset of sample damage. Instead, the diffracted X-rays are gated by a rapid loss of crystalline periodicity, producing apparent pulse lengths significantly shorter than the duration of the incident pulse. The shortest apparent pulse lengths occur at the highest resolution, and our measurements indicate that current X-ray free-electron laser technology5 should enable structural determination from submicrometre protein crystals with atomic resolution.


Structural bases for the specificity of cholinesterase catalysis and inhibition.
The availability of a crystal structure and comparative sequences of the cholinesterases has provided templates suitable for analyzing the molecular bases of specificity of reversible inhibitors, carbamoylating agents and organophosphates. Site-specific mutagenesis enables one to modify the structures of both the binding site and peptide ligand as well as create chimeras reflecting one type of esterase substituted in the template of another. Herein we define the bases for substrate specificity of carboxylesters, the stereospecificity of enantiomeric alkylphosphonates and the selectivity of tricyclic aromatic compounds in the active center of cholinesterase. We also describe the binding loci of the peripheral site and changes in catalytic parameters induced by peripheral site ligands, using the peptide fasciculin.


Birt-Hogg-Dube syndrome is a novel ciliopathy.
Birt-Hogg-Dubé (BHD) syndrome is an autosomal dominant disorder where patients are predisposed to kidney cancer, lung and kidney cysts and benign skin tumors. BHD is caused by heterozygous mutations affecting folliculin (FLCN), a conserved protein that is considered a tumor suppressor. Previous research has uncovered multiple roles for FLCN in cellular physiology, yet it remains unclear how these translate to BHD lesions. Since BHD manifests hallmark characteristics of ciliopathies, we speculated that FLCN might also have a ciliary role. Our data indicate that FLCN localizes to motile and non-motile cilia, centrosomes and the mitotic spindle. Alteration of FLCN levels can cause changes to the onset of ciliogenesis, without abrogating it. In three-dimensional culture, abnormal expression of FLCN disrupts polarized growth of kidney cells and deregulates canonical Wnt signalling. Our findings further suggest that BHD-causing FLCN mutants may retain partial functionality. Thus, several BHD symptoms may be due to abnormal levels of FLCN rather than its complete loss and accordingly, we show expression of mutant FLCN in a BHD-associated renal carcinoma. We propose that BHD is a novel ciliopathy, its symptoms at least partly due to abnormal ciliogenesis and canonical Wnt signalling.


Goodness-of-fit tests for GEE modeling with binary responses.
Analysis of data with repeated measures is often accomplished through the use of generalized estimating equations (GEE) methodology. Although methods exist for assessing the adequacy of the fitted models for uncorrelated data with likelihood methods, it is not appropriate to use these methods for models fitted with GEE methodology. We propose model-based and robust (empirically corrected) goodness-of-fit tests for GEE modeling with binary responses based on partitioning the space of covariates into distinct regions and forming score statistics that are asymptotically distributed as chi-square random variables with the appropriate degrees of freedom. The null distribution and the statistical power of the proposed goodness-of-fit tests were assessed using simulated data. The proposed goodness-of-fit tests are illustrated by two examples using data from clinical studies.


Differential antibody responses to Plasmodium falciparum glycosylphosphatidylinositol anchors in patients with cerebral and mild malaria.
Glycosylphosphatidylinositol (GPI) membrane anchors of Plasmodium falciparum surface proteins are thought to be important factors contributing to malaria pathogenesis, and anti-GPI antibodies have been suggested to provide protection by neutralizing the toxic activity of GPIs. In this study, IgG responses against P. falciparum GPIs and a baculovirus recombinant MSP1p19 antigen were evaluated in two distinct groups of 70 patients each, who were hospitalized with malaria. Anti-GPI IgGs were significantly lower in patients hospitalized with confirmed cerebral malaria compared to those with mild malaria (P < 0.01) but did not discriminate for fatal outcome. In contrast, a specific marker of the anti-parasite immunity, as monitored by the anti-MSP1p19 IgG response, was similar in both cerebral and mild malaria individuals, although it was significantly lower in a subgroup with fatal outcomes. These results are consistent with a potential anti-toxin role for anti-GPI antibodies associated with protection against cerebral malaria.


Simple dialkyl pyrazole-3,5-dicarboxylates show in vitro and in vivo activity against disease-causing trypanosomatids.
The synthesis and antiprotozoal activity of some simple dialkyl pyrazole-3,5-dicarboxylates (compounds 2-6) and their sodium salts (pyrazolates) (compounds 7-9) against Trypanosoma cruzi, Leishmania infantum and Leishmania braziliensis are reported. In most cases the studied compounds showed, especially against the clinically significant amastigote forms, in vitro activities higher than those of the reference drugs (benznidazole for T. cruzi and glucantime for Leishmania spp.); furthermore, the low non-specific cytotoxicities against Vero cells and macrophages shown by these compounds led to good selectivity indexes, which are 8-72 times higher for T. cruzi amastigotes and 15-113 times higher for Leishmania spp. amastigotes than those of the respective reference drugs. The high efficiency of diethyl ester 3 and its sodium salt 8 against the mentioned protozoa was confirmed by further in vitro assays on infection rates and by an additional in vivo study in a murine model of acute and chronic Chagas disease. The inhibitory capacity of compounds 3 and 8 on the essential iron superoxide dismutase of the aforementioned parasites may be related to the observed anti-trypanosomatid activity. The low acute toxicity of compounds 3 and 8 in mice is also reported in this article.


Learning by Demonstration for Motion Planning of Upper-Limb Exoskeletons.
The reference joint position of upper-limb exoskeletons is typically obtained by means of Cartesian motion planners and inverse kinematics algorithms with the inverse Jacobian; this approach allows exploiting the available Degrees of Freedom (i.e. DoFs) of the robot kinematic chain to achieve the desired end-effector pose; however, if used to operate non-redundant exoskeletons, it does not ensure that anthropomorphic criteria are satisfied in the whole human-robot workspace. This paper proposes a motion planning system, based on Learning by Demonstration, for upper-limb exoskeletons that allow successfully assisting patients during Activities of Daily Living (ADLs) in unstructured environment, while ensuring that anthropomorphic criteria are satisfied in the whole human-robot workspace. The motion planning system combines Learning by Demonstration with the computation of Dynamic Motion Primitives and machine learning techniques to construct task- and patient-specific joint trajectories based on the learnt trajectories. System validation was carried out in simulation and in a real setting with a 4-DoF upper-limb exoskeleton, a 5-DoF wrist-hand exoskeleton and four patients with Limb Girdle Muscular Dystrophy. Validation was addressed to (i) compare the performance of the proposed motion planning with traditional methods; (ii) assess the generalization capabilities of the proposed method with respect to the environment variability. Three ADLs were chosen to validate the system: drinking, pouring and lifting a light sphere. The achieved results showed a 100% success rate in the task fulfillment, with a high level of generalization with respect to the environment variability. Moreover, an anthropomorphic configuration of the exoskeleton is always ensured.


Complementation analyses for 45 mutations encompassing the pink-eyed dilution (p) locus of the mouse.
The homozygous and heterozygous phenotypes are described and characterized for 45 new pink-eyed dilution (p) locus mutations, most of them radiation-induced, that affect survival at various stages of mouse development. Cytogenetically detectable aberrations were found in three of the new p mutations (large deletion, inversion, translocation), with band 7C involved in each case. The complementation map developed from the study of 810 types of compound heterozygotes identifies five functional units: jls and jlm (two distinct juvenile-fitness functions, the latter associated with neuromuscular defects), pl-1 and pl-2 (associated with early-postimplantation and preimplantation death, respectively), and nl [neonatal lethality associated with cleft palate (the frequency of rare "escapers" from this defect varied with the genotype)]. Orientation of these units relative to genetic markers is as follows: centromere, Gas-2, pl-1, jls, jlm p, nl (equatable to cp 1 = Gabrb3); pl-2 probably resides in the c-deletion complex. pl-1 does not mask preimplantation lethals between Gas2 and p; and no genes affecting survival are located between p and cp1. The alleles specifying mottling or darker pigment (generically, pm and px, respectively) probably do not represent deletions of p-coding sequences but could be small rearrangements involving proximal regulatory elements.


The layered polyphosphide Ag3.73(4)Zn2.27(4)P16.
The silver zinc hexa-deca-phosphide Ag3.73(4)Zn2.27(4)P16 is the first polyphosphide in the ternary system Ag/Zn/P. It was synthesized from stoichiometric mixtures of Ag, Zn and P in the molar ratio 4:2:16, using AgI as a mineralizing agent in a gas-phase-assisted reaction. Ag3.73(4)Zn2.27(4)P16 crystallizes in the Cu5InP16 structure type. The asymmetric unit contains two Ag/Zn sites with mixed occupancies and four P sites. One of the Ag/Zn sites is located on a twofold rotation axis. The polyanionic [P16]-substructure consists of corrugated six-membered rings that are connected into a layer via the 1-, 2-, 4- and 5-positions of the rings by a bridging P atom in each case. The layers extend parallel to the bc plane and are stacked along the a axis. Both Ag/Zn sites are tetra-hedrally coordinated by P atoms.


LysZX4-NCA, a new endolysin with broad-spectrum antibacterial activity for topical treatment.
The prevalence of multidrug-resistant highly virulent Klebsiella pneumoniae (MDR-hvKP) requires the development of new therapeutic agents. Herein, a novel lytic phage vB_KpnS_ZX4 against MDR-hvKP was discovered in hospital sewage. Phage vB_KpnS_ZX4 had a short latent period (5 min) and a large burst size (230 PFU/cell). It can rapidly reduce the number of bacteria in vitro and improve survival rates of bacteremic mice in vivo from 0 to 80 % with a single injection of 108 PFU. LysZX4, an endolysin derived from vB_KpnS_ZX4, exhibits potent antimicrobial activity in vitro in combination with ethylenediaminetetraacetic acid (EDTA). The antimicrobial activity of LysZX4 was further enhanced by the fusion of KWKLFKI residues from cecropin A (LysZX4-NCA). In vitro antibacterial experiments showed that LysZX4-NCA exerts broad-spectrum antibacterial activity against clinical Gram-negative bacteria, including MDR-hvKP. Moreover, in the mouse model of MDR-hvKP skin infection, treatment with LysZX4-NCA resulted in a three-log reduction in bacterial burden on the skin compared to the control group. Therefore, the novel phages vB_KpnS_ZX4 and LysZX4-NCA are effective reagents for the treatment of systemic and local MDR-hvKP infections.


Can a proton be encapsulated in tetraamido/diamino quaternized macrocycles in aqueous solution and electric field?
The proton-binding behavior of solvated tetraamido/diamino quaternized macrocyclic compounds with rigid phenyl and flexible phenyl bridges in the absence or presence of an external electric field is investigated by molecular dynamics simulation. The proton can be held through H-bonding interactions with the two carbonyl oxygen atoms in macrocycles containing rigid (phenyl) and flexible (propyl) bridges. The solute-solvent H-bonding interactions cause the macrocyclic backbones to twist to different extents, depending on the different bridges. The macrocycle with the rigid phenyl linkages folds into a cuplike shape due to π-π interaction, while the propyl analogue still maintains the ellipsoidal ringlike shape with just a slight distortion. The potential energy required for proton transfer is larger in the phenyl-containing macrocycle than in the compound with propyl units. When an external electric field with a strength of 2.5 V nm(-1) is exerted along the carbonyl oxygen atoms, a difference in proton encircling is exhibited for macrocycles with rigid and flexible bridges. In contrast to encapsulation of a proton in the propyl analogue, the intermolecular solute-solvent H-bonding and intramolecular π-π stacking between the two rigid phenyl spacers leads to loss of the proton from the highly distorted cuplike macrocycle with phenyl bridges. The competition between intra- and intermolecular interactions governs the behavior of proton encircling in macrocycles.


Anti-tumour/metastasis effects of the potassium-sparing diuretic amiloride: an orally active anti-cancer drug waiting for its call-of-duty?
Amiloride.HCl is clinically used as an oral potassium-sparing diuretic, but multiple studies in biochemical, cellular and animal models have shown that the drug also possesses anti-tumour and anti-metastasis activities. The additional effects appear to arise through inhibition of two discrete targets: (i) the sodium-hydrogen exchanger 1 (NHE1), a membrane protein responsible for the characteristically low extracellular pH of tumours and (ii) the urokinase-type plasminogen activator (uPA), a serine protease mediator of cell migration, invasion and metastasis and well-known marker of poor prognosis in cancer. This mini-review summarises for the first time the reported anti-tumour/metastasis effects of amiloride in experimental models, discusses the putative molecular mechanisms responsible for these effects and concludes by commenting on the pros and cons of trialling amiloride or one of its structural analogues as potential new anti-tumour/metastasis drugs.


Risk factors and prognostic factors in patients with double primary cancer: epithelial ovarian cancer and breast cancer.
The most important known risk factor for ovarian cancer is the BRCA1-2 mutation, which is clinically often manifested through a positive family history of cancer of the breast and/or ovary. Whether other risk factors and prognostic factors in women with a positive family history of cancer of the breast and/or ovary and/or with BRCA1-2 mutation are important remains to be elucidated. Recent studies have shown that in the double primary breast and ovarian cancer (DPBOC), BRCA1-2 mutation is present in at least 86% of cases. Therefore, the group of patients with DPBOC, especially with epithelial ovarian cancer and breast cancer, is the most suitable for such an analysis. The aim of this study was to verify the hypothesis that, in this group, some other risk factors, in addition to a specific family history of cancer, as well as unfavourable pathomorphological prognostic factors, are more expressed than in a control group of patients with sporadic epithelial ovarian cancer only. We compared the study group of 31 patients with DPBOC (epithelial ovarian cancer) to a control group of 62 patients with a single, sporadic epithelial ovarian cancer and negative specific family history. The data were obtained from the Cancer Registry of Slovenia and from clinical records. For every patient, we filled-in a protocol and analysed the data, comparing other risk factors in addition to specific family history and prognostic, clinical, and pathomorphological factors. Statistical analysis was performed using descriptive statistics, chi-square test and t-test. Multivariate analysis was also planned, but the necessary conditions were not met. In the study group, we found a higher percentage of positive non-specific family histories than in the control group, but the difference was not statistically significant. No difference in procreative risk factors was observed between the groups. There was a higher percentage of borderline significance of women from the study group that developed ovarian cancer between 45 and 59 years of age. In the study group, ovarian cancer was significantly more often found at Stage I, although the groups did not differ in detection procedures. Also, we did not find any differences in the distribution of tumour grades or histologic tumour types. The results did not confirm our hypothesis, yet they indicated some differences between the groups regarding the risk factors for ovarian cancer. Regarding the prognostic factors, we even found a significantly higher percentage of Stage I epithelial ovarian cancer in the study group, with no difference in the mode of detection. Considering the results that are not typical of BRCA-related cancer (what double primary cancer of the ovary and breast is supposed to be) and previous reports, we find it more likely that the patients with BRCA1-2 mutations represent only a subgroup within the group of patients with double primary breast and ovarian cancer.


Analysis and characterization of hepatitis B vaccine particles synthesized from Hansenula polymorpha.
The biochemical and physical properties of hepatitis B virus (HBV) small surface antigen (S-HBVsAg) from Berna Biotech Korea Corp. were systematically analyzed and characterized. Through various electrophoresis and immunoblotting assay of S-HBVsAg and its proteolytic products, it was confirmed that the S-HBVsAg vaccine particles are present in the form of covalent multimers that are assembled via strong intermolecular disulfide bonds. The S-HBVsAg particles contain no N-glycosylation moiety but some O-glycosidically linked mannoses. Evidently from N-terminus sequencing of both monomers and dimers that are formed by complete and partial reduction, respectively, of the S-HBVsAg particles under reducing SDS-PAGE condition, it is evident that each polypeptide within S-HBVsAg particles has authentic sequence of N-terminus. Denaturation plot shows that the S-HBVsAg vaccine particles were extremely stable especially in the solution with high acidity. This stability property of S-HBVsAg vaccine particles could provide very useful information for the optimization of the downstream process of recombinant S-HBVsAg particles synthesized from yeast cultures.


Fully and partially coherent pathways in multiply enhanced odd-order wave-mixing spectroscopy.
Nuclear magnetic resonance spectroscopy relies on using multiple excitation pulses to create multiple quantum coherences that provide great specificity for chemical measurements. Coherent multidimensional spectroscopy (CMDS) is the optical analogue of NMR. Current CMDS methods use three excitation pulses and phase matching to create zero, single, and double quantum coherences. In order to create higher order multiple quantum coherences, the number of interactions must be increased by raising the excitation intensities high enough to create Rabi frequencies that are comparable to the dephasing rates of vibrational coherences. The higher Rabi frequencies create multiple, odd-order coherence pathways. The coherence pathways that involve intermediate populations are partially coherent and are sensitive to population relaxation effects. Pathways that are fully coherent involve only coherences and measure the direct coupling between excited quantum states. The fully coherent pathways are related to the multiple quantum coherences created in multiple pulse NMR methods such as heteronuclear multiple quantum coherence (HMQC) spectroscopy with the important difference that HMQC NMR methods have a defined number of interactions and avoid dynamic Stark effects whereas the multiply enhanced odd-order wave-mixing pathways do not. The difference arises because CMDS methods use phase matching to define the interactions and at high intensities, multiple pathways obey the same phase matching conditions. The multiple pathways correspond to the pathways created by dynamic Stark effects. This paper uses rhodium dicarbonyl chelate (RDC) as a model to demonstrate the characteristics of multiply enhanced odd-order wave-mixing (MEOW) methods. Dynamic Stark effects excite vibrational ladders on the symmetric and asymmetric CO stretch modes and create a series of multiple quantum coherences and populations using partially and fully coherent pathways. Vibrational quantum states up to v = 6 are excited. A series of spectra provides different two-dimensional cross sections through the multidimensional parameter space involving two excitation frequencies, the frequency of the output coherence, and the excitation pulse time delays. The spectra allow the identification of 18 different overtone and combination band states. Comparison with a local mode model with two anharmonic Morse oscillators with interbond coupling shows excellent agreement.


Multivariate statistical analysis of Raman images of a pharmaceutical tablet.
This paper describes the application of principal component analysis (PCA) and independent component analysis (ICA) to identify the reference spectra of a pharmaceutical tablet's constituent compounds from Raman spectroscopic data. The analysis shows, first with a simulated data set and then with data collected from a pharmaceutical tablet, that both PCA and ICA are able to identify most of the features present in the reference spectra of the constituent compounds. However, the results suggest that the ICA method may be more appropriate when attempting to identify unknown reference spectra from a sample. The resulting PCA and ICA models are subsequently used to estimate the relative concentrations of the constituent compounds and to produce spatial distribution images of the analyzed tablet. These images provide a visual representation of the spatial distribution of the constituent compounds throughout the tablet. Images associated with the ICA scores are found to be more informative and not as affected by measurement noise as the PCA based score images. The paper concludes with a discussion of the future work that needs to be undertaken for ICA to gain wider acceptance in the applied spectroscopy community.


Tobacco addiction as a marker of age at onset of schizophrenia.
Schizophrenia is a heterogenous disorder, with diversity in symptoms, course, prognosis, and probably etiology. The timing of its onset (i.e., early vs. late) not only predicts outcome of illness, but also corresponds to fundamental neurochemical and neuroendocrine distinctions. There is recent evidence that early age at onset of schizophrenia is associated with more prominent negative symptoms, which are associated with decreased dopaminergic functions in the limbic system. Since addictive behaviour may be related to decreased dopamine activity in the mesolimbic reward circuitry, we predict a higher prevalence of tobacco addiction in patients with an earlier age at onset of schizophrenia. To investigate this hypothesis, we studied the association of cigarette smoking to age at onset of illness in a sample of 142 chronic schizophrenic inpatients, 73 of whom were smokers. We found that patients who smoked had a significantly earlier age at onset of psychiatric illness as compared to the nonsmokers (p less than .01). Since damage to dopaminergic systems at the lateral hypothalamic level of the medial forebrain bundle and in the ventral tegmental reward system produces the strongest indication of reward, our data suggest that alterations in dopaminergic functions in these systems may be linked to the timing of onset of schizophrenic symptoms.


Effective "activated PI3Kδ syndrome"-targeted therapy with the PI3Kδ inhibitor leniolisib.
Pathogenic gain-of-function variants in the genes encoding phosphoinositide 3-kinase δ (PI3Kδ) lead to accumulation of transitional B cells and senescent T cells, lymphadenopathy, and immune deficiency (activated PI3Kδ syndrome [APDS]). Knowing the genetic etiology of APDS afforded us the opportunity to explore PI3Kδ inhibition as a precision-medicine therapy. Here, we report in vitro and in vivo effects of inhibiting PI3Kδ in APDS. Treatment with leniolisib (CDZ173), a selective PI3Kδ inhibitor, caused dose-dependent suppression of PI3Kδ pathway hyperactivation (measured as phosphorylation of AKT/S6) in cell lines ectopically expressing APDS-causative p110δ variants and in T-cell blasts derived from patients. A clinical trial with 6 APDS patients was conducted as a 12-week, open-label, multisite, within-subject, dose-escalation study of oral leniolisib to assess safety, pharmacokinetics, and effects on lymphoproliferation and immune dysregulation. Oral leniolisib led to a dose-dependent reduction in PI3K/AKT pathway activity assessed ex vivo and improved immune dysregulation. We observed normalization of circulating transitional and naive B cells, reduction in PD-1+CD4+ and senescent CD57+CD4- T cells, and decreases in elevated serum immunoglobulin M and inflammatory markers including interferon γ, tumor necrosis factor, CXCL13, and CXCL10 with leniolisib therapy. After 12 weeks of treatment, all patients showed amelioration of lymphoproliferation with lymph node sizes and spleen volumes reduced by 39% (mean; range, 26%-57%) and 40% (mean; range, 13%-65%), respectively. Thus, leniolisib was well tolerated and improved laboratory and clinical parameters in APDS, supporting the specific inhibition of PI3Kδ as a promising new targeted therapy in APDS and other diseases characterized by overactivation of the PI3Kδ pathway. This trial was registered at www.clinicaltrials.gov as #NCT02435173.


Translocation of indigenous microflora in an experimental model of sepsis.
Translocation of viable bacteria from gut to bloodstream and other sterile body sites during shock has been demonstrated in several experimental and clinical studies. The factors causing translocation and its incidence at different stages of shock are not known. The aim of the study was to evaluate the importance of several factors causing translocation of indigenous microflora in an experimental model of septic shock based on intraperitoneal Escherichia coli sepsis in rats. Counts of inoculated E. coli and translocated bacteria in different locations, gut morphology and haematological values were evaluated at different stages of sepsis. Sepsis developed in all animals and E. coli achieved the highest counts in blood 6 h after inoculation. Translocation was commonest at 6 and 12 h after inoculation. Frequently translocating bacteria were lactobacilli, bifidobacteria, bacteroides and peptostreptococci. In early sepsis, translocation was associated with high E. coli counts in blood, yet in late sepsis the opposite correlation was present. Low infiltration by neutrophils in the ileum and decreased mitotic activity in the colon were associated with a high translocation rate. In early sepsis, translocation was associated with low lymphocyte counts, but in late sepsis, with low neutrophil counts. Translocation of bacteria (including anaerobes) that colonise the gut in high counts takes place during sepsis. Putative influencing factors such as activity of the primary disease (bacterial counts in blood), gut morphology or haematological values seem to have different impacts on translocation, depending on the stage of the disease.


Bisubstrate UDP-peptide conjugates as human O-GlcNAc transferase inhibitors.
Inhibitors of OGT (O-GlcNAc transferase) are valuable tools to study the cell biology of protein O-GlcNAcylation. We report OGT bisubstrate-linked inhibitors (goblins) in which the acceptor serine in the peptide VTPVSTA is covalently linked to UDP, eliminating the GlcNAc pyranoside ring. Goblin1 co-crystallizes with OGT, revealing an ordered C₃ linker and retained substrate-binding modes, and binds the enzyme with micromolar affinity, inhibiting glycosyltransfer on to protein and peptide substrates.


The dual phosphodiesterase 3/4 inhibitor RPL554 stimulates rare class III and IV CFTR mutants.
Over 2,000 mutations have been reported in the cystic fibrosis transmembrane conductance regulator (cftr) gene, many of which cause disease but are rare and have no effective treatment. Thus, there is an unmet need for new, mutation-agnostic therapies for cystic fibrosis (CF). Phosphodiesterase (PDE) inhibitors are one such class of therapeutics that have been shown to elevate intracellular cAMP levels and stimulate CFTR-dependent anion secretion in human airway epithelia; however, the number of people with CF that could be helped by PDE inhibitors remains to be determined. Here we used Fisher rat thyroid (FRT) cells stably transduced with rare human CFTR mutants and studied their responsiveness to the dual phosphodiesterase 3/4 inhibitor RPL554 (Verona Pharma). Through its inhibitory effect on PDE4D, we find that RPL554 can elevate intracellular cAMP leading to a potentiation of forskolin-stimulated current mediated by R334W, T338I, G551D, and S549R mutants of CFTR when used alone or in combination with CFTR modulators. We also were able to reproduce these effects of RPL554 on G551D-CFTR when it was expressed in primary human bronchial epithelial cells, indicating that RPL554 would have stimulatory effects on rare CFTR mutants in human airways and validating FRT cells as a model for PDE inhibitor studies. Furthermore, we provide biochemical evidence that VX-809 causes surprisingly robust correction of several class III and IV CFTR mutants. Together, our findings further support the therapeutic potential of RPL554 for patients with CF with class III/IV mutations and emphasize the potential of PDEs as potential drug targets that could benefit patients with CF.


Integrated microfluidic pumps and valves operated by finger actuation.
Here, we report an integrated operation of microfluidic pumps and valves only by finger actuation. As the working principle of the finger-actuated microfluidic pumps includes deflection of the poly(dimethylsiloxane) (PDMS) membrane, the pneumatic valves for controlling the flow direction can be easily integrated with the pumps. Using a single button, the flow path can be determined and flow generation can be achieved. We also verified the integrated operation of finger-actuated pumps and valves by demonstrating nucleic acid purification.


Neuronal MT1-MMP mediates ECM clearance and Lrp4 cleavage for agrin deposition and signaling in presynaptic development.
Agrin is a crucial factor that induces postsynaptic differentiation at neuromuscular junctions (NMJs), but how secreted agrin is locally deposited in the context of extracellular matrix (ECM) environment and its function in presynaptic differentiation remain largely unclear. Here, we report that the proteolytic activity of neuronal membrane-type 1 matrix metalloproteinase (MT1-MMP; also known as MMP14) facilitates agrin deposition and signaling during presynaptic development at NMJs. Firstly, agrin deposition along axons exhibits a time-dependent increase in cultured neurons that requires MMP-mediated focal ECM degradation. Next, local agrin stimulation induces the clustering of mitochondria and synaptic vesicles, two well-known presynaptic markers, and regulates vesicular trafficking and surface insertion of MT1-MMP. MMP inhibitor or MT1-MMP knockdown suppresses agrin-induced presynaptic differentiation, which can be rescued by treatment with the ectodomain of low-density lipoprotein receptor-related protein 4 (Lrp4). Finally, neuronal MT1-MMP knockdown inhibits agrin deposition and nerve-induced acetylcholine receptor clustering in nerve-muscle co-cultures and affects synaptic structures at Xenopus NMJs in vivo Collectively, our results demonstrate a previously unappreciated role of agrin, as well as dual functions of neuronal MT1-MMP proteolytic activity in orchestrating agrin deposition and signaling, in presynaptic development.


Recent advances in the development and use of molecular tests to predict antimicrobial resistance in Neisseria gonorrhoeae.
The number of genetic tests, mostly real-time PCRs, to detect antimicrobial resistance (AMR) determinants and predict AMR in Neisseria gonorrhoeae is increasing. Several of these assays are promising, but there are important shortcomings and few assays have been adequately validated and quality assured. Areas covered: Recent advances, focusing on publications since 2012, in the development and use of molecular tests to predict gonococcal AMR for surveillance and for clinical use, advantages and disadvantages of these tests and of molecular AMR prediction compared with phenotypic AMR testing, and future perspectives for effective use of molecular AMR tests for different purposes. Expert commentary: Several challenges for direct testing of clinical, especially extra-genital, specimens remain. The choice of molecular assay needs to consider the assay target, quality controls, sample types, limitations intrinsic to molecular technologies, and specific to the chosen methodology, and the intended use of the test. Improved molecular- and particularly genome-sequencing-based methods will supplement AMR testing for surveillance purposes, and translate into point-of-care tests that will lead to personalized treatments, while sparing the last available empiric treatment option (ceftriaxone). However, genetic AMR prediction will never completely replace phenotypic AMR testing, which detects also AMR due to unknown AMR determinants.


Association Between Borderline Personality Features and Temporal Summation of Second Pain: A Cross-Sectional Study.
Individuals with greater borderline personality features may be vulnerable to chronic pain. Because pain is an unpleasant sensory and emotional experience, affect dysregulation as the core personality feature may be linked to pain hypersensitivity. Studies have found that greater borderline features are associated with increased intensity in clinical and experimental pain, and that depression mediates this increase. The current study further examined the association between borderline features and heat pain sensitivity, the contribution of affect dysregulation and the other borderline personality factors (identity problems, negative relationships, self-harming/impulsivity) to the association, and depression as a mediator. Additionally, we examined whether blunted sympathetic responses mediate the association between borderline features and temporal summation of second pain (TSSP). Thermal pain threshold, thermal TSSP and aftersensations pain were assessed in 79 healthy individuals with varying degrees of borderline features. TSSP is a proxy measure for central sensitization and refers to the gradual increase in pain to repeated nociceptive stimuli. A regression analysis showed that greater borderline features predicted greater TSSP (β = .22, p = .050, R2 = .05). Borderline features were unrelated to pain threshold and TSSP decay. A stepwise regression showed greater TSSP in individuals with greater borderline features was accounted for by the negative relationships factor rather than the affect dysregulation factor. The results of mediational analyses showed depression and blunted sympathetic skin conductance responses mediated the positive association between TSSP and borderline features.


BiosyntheticSPAdes: reconstructing biosynthetic gene clusters from assembly graphs.
Predicting biosynthetic gene clusters (BGCs) is critically important for discovery of antibiotics and other natural products. While BGC prediction from complete genomes is a well-studied problem, predicting BGCs in fragmented genomic assemblies remains challenging. The existing BGC prediction tools often assume that each BGC is encoded within a single contig in the genome assembly, a condition that is violated for most sequenced microbial genomes where BGCs are often scattered through several contigs, making it difficult to reconstruct them. The situation is even more severe in shotgun metagenomics, where the contigs are often short, and the existing tools fail to predict a large fraction of long BGCs. While it is difficult to assemble BGCs in a single contig, the structure of the genome assembly graph often provides clues on how to combine multiple contigs into segments encoding long BGCs. We describe biosyntheticSPAdes, a tool for predicting BGCs in assembly graphs and demonstrate that it greatly improves the reconstruction of BGCs from genomic and metagenomics data sets.


Enhancement of gene expression efficiency using cationic liposomes on ovarian cancer cells.
We performed transfection using cationic liposome. According to the gene expression level, lined cells were divided into two groups, high and low. Introduced gene was monitored with a confocal laser-scanning microscope. The percentages of the cells introduced gene reached more than 90% in all line. Then, introduced gene was stable in high group, while in low group, it significantly decreased. With lysosomotrophic agents, gene expression efficiency was significantly reduced.With colchicine, gene expression efficiency did not change in high group, but was significantly elevated in low group. A method of liposomal transfection could be effective, particularly in low group.


Prognostic value of B-type natriuretic peptide in elderly patients with aortic valve stenosis: the COFRASA-GENERAC study.
Previous studies suggested an independent prognostic value of B-type natriuretic peptide (BNP) in aortic valve stenosis (AS) but were impeded by small sample sizes and inclusion of relatively selected young patients. We aimed to evaluate the relationship among N-terminal fragment of proBNP (Nt-proBNP), AS severity, symptoms and outcome in a large cohort of elderly patients with AS. Observational cohort study, COhorte Française de Retrecissement Aortique du Sujet Agé (clinicalTrial.gov number-NCT00338676) and GENEtique du Retrecissement Aortique (clinicalTrial.gov number-NCT00647088). Single-centre study. Patients older than 70 years with at least mild AS. None. A comprehensive clinical, biological and echocardiographic evaluation was performed at study entry. Asymptomatic patients were prospectively followed on a 6-months basis and AS-related events (sudden death, congestive heart failure or new onset of AS-related symptoms) collected. We prospectively enrolled 361 patients (79±6 years, 230 severe AS). Nt-proBNP increased with the grade of AS severity and the NYHA class (all p<0.0001) but there was an important overlap between grades/classes. Consequently, diagnostic value of Nt-proBNP for the diagnosis of severe symptomatic AS was only modest (area under the curve of the receiver operator characteristic analysis=0.73). At 2 years, 28 AS-related events occurred among 142 asymptomatic patients prospectively followed. Nt-proBNP was associated with outcome in univariate analysis (p=0.04) but not after adjustment for age, gender and AS severity (p=0.40). The present study clearly highlights the limitations of Nt-proBNP for the evaluation and management of AS patients. Our results suggest that Nt-proBNP should be considered cautiously, at least as a single criterion, in the decision-making process of AS patients especially in the elderly population.


Identification of mycobactins by nuclear-magnetic-resonance spectroscopy.
Details are given of the n.m.r. spectra of mycobactins F, H, M, N, P, S and T, and resonances are ascribed to all the protons in these molecules. A simplified system is described for identifying known mycobactins by the n.m.r. spectrum alone. This method will not distinguish mycobactins S and T, whose nuclei differ only in the configuration at an asymmetric centre.


Molecular pathways: targeted α-particle radiation therapy.
An α-particle, a (4)He nucleus, is exquisitely cytotoxic and indifferent to many limitations associated with conventional chemo- and radiotherapy. The exquisite cytotoxicity of α-radiation, the result of its high mean energy deposition [high linear energy transfer (LET)] and limited range in tissue, provides for a highly controlled therapeutic modality that can be targeted to selected malignant cells [targeted α-therapy (TAT)] with minimal normal tissue effects. A burgeoning interest in the development of TAT is buoyed by the increasing number of ongoing clinical trials worldwide. The short path length renders α-emitters suitable for treatment and management of minimal disease such as micrometastases or residual tumor after surgical debulking, hematologic cancers, infections, and compartmental cancers such as ovarian cancer or neoplastic meningitis. Yet, despite decades of study of high LET radiation, the mechanistic pathways of the effects of this modality remain not well defined. The modality is effectively presumed to follow a simple therapeutic mechanism centered on catastrophic double-strand DNA breaks without full examination of the actual molecular pathways and targets that are activated that directly affect cell survival or death. This Molecular Pathways article provides an overview of the mechanisms and pathways that are involved in the response to and repair of TAT-induced DNA damage as currently understood. Finally, this article highlights the current state of clinical translation of TAT as well as other high-LET radionuclide radiation therapy using α-emitters such as (225)Ac, (211)At, (213)Bi, (212)Pb, and (223)Ra.


Distinct recruitment of dorsomedial and dorsolateral striatum erodes with extended training.
Hypotheses of striatal orchestration of behavior ascribe distinct functions to striatal subregions, with the dorsolateral striatum (DLS) especially implicated in habitual and skilled performance. Thus neural activity patterns recorded from the DLS, but not the dorsomedial striatum (DMS), should be correlated with habitual and automatized performance. Here, we recorded DMS and DLS neural activity in rats during training in a task promoting habitual lever pressing. Despite improving performance across sessions, clear changes in corresponding neural activity patterns were not evident in DMS or DLS during early training. Although DMS and DLS activity patterns were distinct during early training, their activity was similar following extended training. Finally, performance after extended training was not associated with DMS disengagement, as would be predicted from prior work. These results suggest that behavioral sequences may continue to engage both striatal regions long after initial acquisition, when skilled performance is consolidated.


Investigation of an epidemic of acute haemorrhagic conjunctivitis in Pune, India.
An epidemic of acute haemorrhagic conjunctivitis (AHC) was reported from Pune in August-September, 1996. Conjunctival swabs and scrapings were taken from acute patients attending the OPD of the Ophthalmology department of B. J. Medical College, Pune. Conjunctival swabs were processed in human embryonic lung fibroblasts (HEL) cell line and viral isolates were obtained which were identified as Enterovirus (EV) 70 prime strain using standard in-vitro neutralisation test. Indirect intracellular immunofluorescence test also confirmed the identity of the virus involved in the epidemic.


Nutrient sensing and signalling by the gut.
Recent advances highlight that nutrient receptors (such as T1R1/T1R3 heterodimer, Ca sensing receptor and GPR93 for amino acids and protein, GPR40, GPR41, GPR43 and GPR120 for fatty acids, T1R2/T1R3 heterodimer for monosaccharides) are expressed in the apical face of the gut and sense nutrients in the lumen. They transduce signals for the regulation of nutrient transporter expressions in the apical face. Interestingly, they are also localised in enteroendocrine cells (EEC) and mainly exert a direct control on the secretion in the lamina propria of gastro-intestinal peptides such as cholecystokinin, glucagon-like peptide-1 and peptide YY in response to energy nutrient transit and absorption in the gut. This informs central nuclei involved in the control of feeding such as the hypothalamus and nucleus of the solitary tract of the availability of these nutrients and thus triggers adaptive responses to maintain energy homoeostasis. These nutrient receptors then have a prominent position since they manage nutrient absorption and are principally the generator of the first signal of satiation mechanisms mainly transmitted to the brain by vagal afferents. Moreover, tastants are also able to elicit gut peptides secretion via chemosensory receptors expressed in EEC. Targeting these nutrient and tastant receptors in EEC may thus be helpful to promote satiation and so to fight overfeeding and its consequences.


Phase behaviour of liquid-crystal monolayers of rod-like and plate-like particles.
Orientational and positional ordering properties of liquid crystal monolayers are examined by means of Fundamental-Measure Density Functional Theory. Particles forming the monolayer are modeled as hard parallelepipeds of square section of size σ and length L. Their shapes are controlled by the aspect ratio κ = L/σ (>1 for prolate and <1 for oblate shapes). The particle centers of mass are restricted to a flat surface and three possible and mutually perpendicular orientations (in-plane and along the layer normal) of their uniaxial axes are allowed. We find that the structure of the monolayer depends strongly on particle shape and density. In the case of rod-like shapes, particles align along the layer normal in order to achieve the lowest possible occupied area per particle. This phase is a uniaxial nematic even at very low densities. In contrast, for plate-like particles, the lowest occupied area can be achieved by random in-plane ordering in the monolayer, i.e., planar nematic ordering takes place even at vanishing densities. It is found that the random in-plane ordering is not favorable at higher densities and the system undergoes an in-plane ordering transition forming a biaxial nematic phase or crystallizes. For certain values of the aspect ratio, the uniaxial-biaxial nematic phase transition is observed for both rod-like and plate-like shapes. The stability region of the biaxial nematic phase enhances with decreasing aspect ratios for plate-like particles, while the rod-like particles exhibit a reentrant phenomenon, i.e., a sequence of uniaxial-biaxial-uniaxial nematic ordering with increasing density if the aspect ratio is larger than 21.34. In addition to this, packing fraction inversion is observed with increasing surface pressure due to the alignment along the layers normal. At very high densities the nematic phase destabilizes to a nonuniform phases (columnar, smectic, or crystalline phases) for both shapes.


Endogenous thrombin potential in Behçet's disease: relationship with thrombosis and anticoagulant therapy.
To analyse the relationship between an automated thrombin generation test, the endogenous thrombin potential (ETP), and other hypercoagulability markers, with vascular involvement in patients with Behçet's disease (BD). Patients and methods. We analysed 56 BD patients (30 men; mean age, 34.4 ± 14.3 years) without any known thrombophilic factor, of which 17 had previously suffered from thrombosis (deep venous thrombosis in 14 and ischaemic stroke in 3), and 56 controls matched for age and sex. Additionally, we also evaluated 20 plasma samples with an international normalised ratio (INR) between 1.5 and 5.0 obtained from patients with atrial fibrillation but without a history of embolic events that were under treatment with acenocumarol. Thrombin generation was measured as ETP with a chromogenic assay in an automated analyser. Factor VIII, von Willebrand factor antigen, prothrombin fragment 1.2, D-dimer and plasmin-antiplasmin complexes were also measured. BD patients showed higher ETP values than controls (471.3± 49.3 vs. 427.5± 31.3 mA; p<0.001). Additionally, BD patients with a history of thrombosis had higher ETP values than patients without thrombosis (496.6± 36.5 vs. 460.7± 50.5 mA; p<0.01). Factor VIII and von Willebrand factor antigen were also elevated in BD patients, but only von Willebrand factor antigen showed statistically significant differences between BD patients with and without thrombosis. Acenocumarol treatment reduced thrombin generation in BD patients in parallel to INR levels, reaching values similar to those of patients with atrial fibrillation and similar INR. BD is associated with thrombosis, and increased thrombin generation (measured as ETP) is a promising marker of hypercoagulability.


Tinnitus Suppression by Intracochlear Electrical Stimulation in Single Sided Deafness - A Prospective Clinical Trial: Follow-Up.
Earlier studies show that a Cochlear Implant (CI), capable of providing intracochlear electrical stimulation independent of environmental sounds, appears to suppress tinnitus at least for minutes. The current main objective is to compare the long-term suppressive effects of looped (i.e. repeated) electrical stimulation (without environmental sound perception) with the standard stimulation pattern of a CI (with environmental sound perception). This could open new possibilities for the development of a "Tinnitus Implant" (TI), an intracochlear pulse generator for the suppression of tinnitus. Ten patients with single sided deafness suffering from unilateral tinnitus in the deaf ear are fitted with a CI (MED-EL Corporation, Innsbruck, Austria). Stimulation patterns are optimized for each individual patient, after which they are compared using a randomized crossover design, with a follow-up of six months, followed by a 3 month period using the modality of patient's choice. Results show that tinnitus can be suppressed with intracochlear electrical stimulation independent of environmental sounds, even long term. No significant difference in tinnitus suppression was found between the standard clinical CI and the TI. It can be concluded that coding of environmental sounds is no requirement for tinnitus suppression with intracochlear electrical stimulation. It is therefore plausible that tinnitus suppression by CI is not solely caused by an attention shift from the tinnitus to environmental sounds. Both the standard clinical CI and the experimental TI are potential treatment options for tinnitus. These findings offer perspectives for a successful clinical application of the TI, possibly even in patients with significant residual hearing. TrialRegister.nl NTR3374.


The use of cardiovascular risk factor information in practice databases: making the best of patient data.
Primary care teams record cardiovascular risk factor data on their patients to help them identify and treat patients eligible for prevention. However, it is not known to what extent this information is already available to clinicians, or the extent to which it is used. To assess the extent to which risk factor is recorded, and to determine the cost-effectiveness of using recorded risk factor information in order to identify and treat eligible patients. An Excel-based model of the incremental costs and benefits of assessment and treatment. Two general practices in the West Midlands. Untreated, non-diabetic patients, aged 35-74 years, were identified from each practice, and risk factor data was uploaded into an Excel spreadsheet. The completeness of risk factor data was assessed. The costs and benefits of assessing and treating patients, in descending order of estimated cardiovascular risk, were then modelled. In each practice, 72.9% and 77.7% of patients had a record of their blood pressure, 26.9% and 25.7% were eligible for at least one treatment: aspirin was the most common treatment followed by antihypertensives. With patients systematically assessed in descending order of cardiovascular risk, 78% of eligible patients and 87% of preventable cardiovascular events are found in the first two deciles of the target population. Lack of risk factor information is not the principal constraint on cardiovascular prevention. Practices have sufficient risk factor data to inform an efficient, targeted prevention strategy.


Integrated network pharmacology and targeted metabolomics to reveal the mechanism of nephrotoxicity of triptolide.
Triptolide (TP) is one of the important active components in Tripterygium wilfordii Hook. F., which shows strong anti-inflammatory and immunomodulatory effects. However, a large number of literature studies have reported that TP is the main component causing nephrotoxicity, and the mechanism of nephrotoxicity has not yet been revealed. Therefore, it is of great practical significance to clarify the toxicity mechanism of TP. This study integrated network pharmacology and targeted metabolomics to reveal the nephrotoxicity mechanism of TP. Firstly, network pharmacology screening of 61 action targets related to TP induced nephrotoxicity, with 39 direct targets and 22 indirect targets, was performed. Subsequently, based on a large-scale protein-protein interaction (PPI) and molecular docking validation, the core targets were identified. Based on the above targets and enrichment analysis, the purine metabolism, Toll-like receptor signaling pathway and NF-κB signaling pathway were found play a pivotal role in TP-induced nephrotoxicity. Literature investigation showed that purine and pyrimidine metabolism pathways were closely related to kidney diseases. Therefore, by using the quantitative method of determining endogenous purine and pyrimidine previously established in the laboratory, a targeted metabolomic analysis of TP was carried out. Finally, six nephrotoxicity biomarkers, dihydroorotate, thymidine, 2-deoxyinosine, uric acid, adenosine and xanthine, were found. Combining the above results, the mechanisms underlying the nephrotoxicity of TP were speculated to be due to the over-consumption of xanthine and uric acid, which would result in enormous ROS being released in response to oxidative stress in the body. Furthermore, activation of the Toll-like receptor signalling pathway can promotes the phosphorylation of the downstream protein NF-κB and causes an inflammatory response that ultimately leads to nephrotoxicity.


Evaluation of an Innovative Case Management Strategy to Improve Rehabilitation Coverage for People at High Risk of Permanent Work Disability: a Feasibility Study.
Our process evaluation (trial registration: DRKS00022468) monitored the implementation of a multi-component strategy including case management aiming to improve rehabilitation coverage and work participation of people with a high risk of permanent work disability. A risk score using administrative data, particularly on employment and welfare benefits, was employed to identify individuals with higher probability of receiving disability pension and therefore potentially needing support. These individuals were contacted by post and encouraged to phone their regional case manager if they needed assistance. Content for the intervention components was developed collaboratively with the case managers. We examined the sample reached, dose delivered, fidelity, dose received and satisfaction with the intervention. Out of 1074 individuals with high-risk scores were contacted, there were 57 case managements. The participants reached were in poor health, and 42.1% reported at least four diagnosed conditions, mostly musculoskeletal and mental disorders. About two-thirds (63.0%) reported poor work ability at baseline. On average, 72.5% of the content of the initial telephone contacts, 88.7% of the content of the face-to-face interviews and 45.2% of the content of the case management were delivered. The participants were highly satisfied with the various components and content of the intervention. Knowledge about rehabilitation improved significantly, with 43 of the 57 participants (75.4%) applying for rehabilitation, mainly medical rehabilitation. At the end of the case management intervention, most participants (91.7%) were still employed. The risk score offers an opportunity to screen for people with a high risk of permanent work disability. Case management participants found participation worthwhile and were significantly better informed about participation services after completing case management. More than every second participant received medical or vocational rehabilitation.


Regulation of regional vascular tone: the role of angiotensin conversion in human forearm resistance vessels.
Inhibition of the action of endothelially-located angiotensin converting enzyme (ACE) in blood vessels of the human forearm was studied using enalaprilat, the active metabolite of the prodrug enalapril. In a dose of 5 micrograms/min enalaprilat inhibits arteriolar vasoconstriction in response to angiotensin I (Ang I) and enhances vasodilation in response to bradykinin. At this dose enalaprilat had no effect on resting forearm blood flow, or on the reduction in forearm blood flow in response to application of lower body negative pressure, in subjects with normal sodium intake. Following sodium depletion, however, enalaprilat produced an increase in resting forearm blood flow compared with the response in the same subjects under normal-sodium conditions. It appears that local ACE within forearm resistance vessels of healthy volunteers is unlikely to play an important role in regulation of local vascular tone in the sodium-replete state. However, in sodium-depleted subjects, and perhaps also in other circumstances where circulating concentrations of Ang I are elevated, local ACE may significantly affect vascular tone.


Empagliflozin Improves the MicroRNA Signature of Endothelial Dysfunction in Patients with Heart Failure with Preserved Ejection Fraction and Diabetes.
Endothelial dysfunction represents a key mechanism underlying heart failure with preserved ejection fraction (HFpEF), diabetes mellitus (DM), and frailty. However, reliable biomarkers to monitor endothelial dysfunction in these patients are lacking. In this study, we evaluated the expression of a panel of circulating microRNAs (miRs) involved in the regulation of endothelial function in a population of frail older adults with HFpEF and DM treated for 3 months with empagliflozin, metformin, or insulin. We identified a distinctive pattern of miRs that were significantly regulated in HFpEF patients compared to healthy controls and to HFpEF patients treated with the sodium glucose cotransporter 2 (SGLT2) inhibitor empagliflozin. Three miRs were significantly downregulated (miR-126, miR-342-3p, and miR-638) and two were significantly upregulated (miR-21 and miR-92) in HFpEF patients compared to healthy controls. Strikingly, two of these miRs (miR-21 and miR-92) were significantly reduced in HFpEF patients after the 3-month treatment with empagliflozin, whereas no significant differences in the profile of endothelial miRs were detected in patients treated with metformin or insulin. Taken together, our findings demonstrate for the first time that specific circulating miRs involved in the regulation of endothelial function are significantly regulated in frail HFpEF patients with DM and in response to SGLT2 inhibition. SIGNIFICANCE STATEMENT: We have identified a novel microRNA signature functionally involved in the regulation of endothelial function that is significantly regulated in frail patients with HFpEF and diabetes. Moreover, the treatment with the SGLT2 inhibitor empagliflozin caused a modification of some of these microRNAs in a direction that was opposite to what observed in HFpEF patients, indicating a rescue of endothelial function. Our findings are relevant for clinical practice inasmuch as we were able to establish novel biomarkers of disease and response to therapy.


Helicobacter pylori-induced peptic ulcer disease is associated with inadequate regulatory T cell responses.
Helicobacter pylori infection is the major cause of peptic ulceration and gastric adenocarcinoma. To address the hypothesis that the human acquired immune response to H. pylori influences pathogenesis, we characterised the gastric T helper (Th) and regulatory T cell (Treg) response of infected patients. The human gastric CD4(+) T cell response of 28 donors who were infected with H. pylori and 44 who were not infected was analysed using flow cytometry. The T cell associated mucosal cytokine response was analysed by real-time polymerase chain reaction assay of samples from 38 infected and 22 uninfected donors. Recombinant interleukin 10 (IL10) was added to co-cultures of H. pylori and AGS cells and its suppressive effects upon inflammatory responses were measured. We found that the H. pylori-specific response consists of both T helper 1 and 2 subsets with high levels of IL10-secreting Tregs. People with peptic ulcer disease had a 2.4-fold reduced CD4(+)CD25(hi)IL10(+) Treg response (p = 0.05) but increased Th1 and Th2 responses (Th1: 3.2-fold, p = 0.038; Th2: 6.1-fold, p = 0.029) compared to those without ulcers. In vitro studies showed that IL10 inhibited IL8 expression and activation of nuclear factor kappa B induced by H. pylori in gastric epithelial cells, and enhanced H. pylori growth in a bacterial-cell co-culture model. Together our data suggest that H. pylori induces a regulatory T cell response, possibly contributing to its peaceful coexistence with the human host, and that ulcers occur when this regulatory response is inadequate.


Comparison of olecranon plate fixation in osteoporotic bone: do current technologies and designs make a difference?
The purpose of this study is to determine if recent innovations in olecranon plates have any advantages in stabilizing osteoporotic olecranon fractures. Five olecranon plates (Acumed, Synthes-SS, Synthes-Ti, US Implants/ITS, and Zimmer) were implanted to stabilize a simulated comminuted fracture pattern in 30 osteoporotic cadaveric elbows. Specimens were randomized by bone mineral density per dual-energy x-ray absorptiometry scan. Three-dimensional displacement analysis was conducted to assess fragment motion through physiological cyclic arcs of motion and failure loading, which was statistically compared using one-way analysis of variance and Tukey honestly significant difference post hoc comparisons with a critical significance level of α = 0.05. Bone mineral density ranged from 0.546 g/cm to 0.878 g/cm with an average of 0.666 g/cm. All implants limited displacement of the fragments to less than 3 mm until sudden, catastrophic failure as the bone of the proximal fragment pulled away from the implant. The maximum load sustained by all osteoporotic specimens ranged from 1.6 kg to 6.6 kg with an average of 4.4 kg. There was no statistical difference between the groups in terms of cycles survived and maximum loads sustained. Cyclic physiological loading of osteoporotic olecranon fracture fixation resulted in sudden, catastrophic failure of the bone-implant interface rather than in gradual implant loosening. Recent plate innovations such as locking plates and different screw designs and positions appear to offer no advantages in stabilizing osteoporotic olecranon fractures. Surgeons may be reassured that the current olecranon plates will probably adequately stabilize osteoporotic fractures for early motion in the early postoperative period, but not for heavy activities such as those that involve over 4 kg of resistance.


Comparison of maternity experiences of Canadian-born and recent and non-recent immigrant women: findings from the Canadian Maternity Experiences Survey.
To compare the maternity experiences of immigrant women (recent, ≤ 5 years in Canada; non-recent > 5 years) with those of Canadian-born women. This study was based on data from the Canadian Maternity Experiences Survey of the Public Health Agency of Canada. A stratified random sample of 6421 women was drawn from a sampling frame based on the 2006 Canadian Census of Population. Weighted proportions were calculated using survey sample weights. Multivariable logistic regression was used to estimate odds ratios comparing recent immigrant women with Canadian-born women and non-recent immigrant women with Canadian-born women, adjusting for education, income, parity, and maternal age. The sample comprised 7.5% recent immigrants, 16.3% non-recent immigrants, and 76.2% Canadian-born women. Immigrant women reported experiencing less physical abuse and stress, and they were less likely to smoke or consume alcohol during pregnancy, than Canadian women; however, they were more likely to report high levels of postpartum depression symptoms and were less likely to have access to social support, to take folic acid before and during pregnancy, to rate their own and their infant's health as optimal, and to place their infants on their backs for sleeping. Recent and non-recent immigrant women also had different experiences, suggesting that duration of residence in Canada plays a role in immigrant women's maternity experiences. These findings can assist clinicians and policy-makers to understand the disparities that exist between immigrant and non-immigrant women in order to address the needs of immigrant women more effectively.


The Nature of Finger Enslaving: New Results and Their Implications.
We present a review on the phenomenon of unintentional finger action seen when other fingers of the hand act intentionally. This phenomenon (enslaving) has been viewed as a consequence of both peripheral (e.g., connective tissue links and multifinger muscles) and neural (e.g., projections of corticospinal pathways) factors. Recent studies have shown relatively large and fast drifts in enslaving toward higher magnitudes, which are not perceived by subjects. These and other results emphasize the defining role of neural factors in enslaving. We analyze enslaving within the framework of the theory of motor control with spatial referent coordinates. This analysis suggests that unintentional finger force changes result from drifts of referent coordinates, possibly reflecting the spread of cortical excitation.


Spectral heterogeneity in single light-harvesting chlorosomes from green sulfur photosynthetic bacterium chlorobium tepidum.
The fluorescence emission properties of single chlorosomes from the green sulfur photosynthetic bacterium Chlorobium (Chl.) tepidum are studied for the first time, using a total internal reflection fluorescence microscope. The fluorescence peak positions of bacteriochlorophyll (BChl)-c self-aggregates in a single chlorosome of Chl. tepidum were widely distributed in the wavelength region between 750 and 768 nm, and the standard deviation (s.d. = 4.1 nm, n = 51) was larger than that of single chlorosomes of Chloroflexus (Cfl.) (s.d. = 1.9 nm, n = 50). The spectral heterogeneity among single chlorosomes from Chl. tepidum was in sharp contrast to those from Cfl. aurantiacus. The difference of chlorosomal spectral properties between Chl. tepidum and Cfl. aurantiacus at the single-unit level would be ascribed to the homolog composition of BChl-c--chlorosomes of Chl. tepidum have BChl substituted with various alkyl groups at both the 8- and 12-positions, whereas light-harvesting BChl-c molecules in Cfl. chlorosomes have the same substituents at the 8- (ethyl group) and 12- (methyl group) positions.


Volunteer studies investigating the safety and efficacy of live oral El Tor Vibrio cholerae O1 vaccine strain CVD 111.
A live oral cholera vaccine should ideally protect against both classical and El Tor biotypes of Vibrio cholerae O1. An El Tor biotype vaccine strain, therefore, would complement classical cholera vaccine strain CVD 103-HgR, a strain already in use in some countries. In this study, 25 healthy adult volunteers received a single dose of 10s colony-forming units of El Tor vaccine strain CVD 111, a derivative of El Tor Ogawa strain N16117 deleted in the virulence cassette. Three (12%) volunteers developed mild diarrhea (mean stool volume = 813 ml) but no systemic symptoms; 23 (92%) of the 25 volunteers developed serum vibriocidal antibodies (geometric mean titer = 1:2,291). Five weeks after vaccination, 18 vaccines and eight uninimunized control volunteers underwent wild-type challenge with El Tor Ogawa strain 3008. Three (16.7%) of 18 vaccinees and seven (87.5%) of eight controls developed diarrhea (P = 0.001) (vaccine efficacy = 80.9%). Further studies are underway to determine a dosage of CVD 111 that will be more clinically acceptable but equally immunogenic and protective.


Cardiac myofibrillar contractile properties during the progression from hypertension to decompensated heart failure.
Heart failure arises, in part, from a constellation of changes in cardiac myocytes including remodeling, energetics, Ca2+ handling, and myofibrillar function. However, little is known about the changes in myofibrillar contractile properties during the progression from hypertension to decompensated heart failure. The aim of the present study was to provide a comprehensive assessment of myofibrillar functional properties from health to heart disease. A rodent model of uncontrolled hypertension was used to test the hypothesis that myocytes in compensated hearts exhibit increased force, higher rates of force development, faster loaded shortening, and greater power output; however, with progression to overt heart failure, we predicted marked depression in these contractile properties. We assessed contractile properties in skinned cardiac myocyte preparations from left ventricles of Wistar-Kyoto control rats and spontaneous hypertensive heart failure (SHHF) rats at ~3, ~12, and >20 mo of age to evaluate the time course of myofilament properties associated with normal aging processes compared with myofilaments from rats with a predisposition to heart failure. In control rats, the myofilament contractile properties were virtually unchanged throughout the aging process. Conversely, in SHHF rats, the rate of force development, loaded shortening velocity, and power all increased at ~12 mo and then significantly fell at the >20-mo time point, which coincided with a decrease in left ventricular fractional shortening. Furthermore, these changes occurred independent of changes in β-myosin heavy chain but were associated with depressed phosphorylation of myofibrillar proteins, and the fall in loaded shortening and peak power output corresponded with the onset of clinical signs of heart failure.NEW & NOTEWORTHY This novel study systematically examined the power-generating capacity of cardiac myofilaments during the progression from hypertension to heart disease. Previously undiscovered changes in myofibrillar power output were found and were associated with alterations in myofilament proteins, providing potential new targets to exploit for improved ventricular pump function in heart failure.


Innovative lead compounds and formulation strategies as newer kinetoplastid therapies.
The protozoan diseases leishmaniasis, human African trypanosomiasis (HAT) and Chagas disease (CD) are responsible for substantial global morbidity and mortality in tropical and subtropical regions. Environmental changes, drug resistance and immunosuppression are contributing to the emergence and spread of these diseases. In the absence of safe and efficient vaccines, chemotherapy, together with vector control, remains the most important measure to control kinetoplastid diseases. Nevertheless, the current chemotherapeutic treatments are clearly inadequate because of their toxic effects, generation of resistances as well as route and schedules of administration not adapted to the field-conditions. This review overlooks the strategies that can be addressed to meet immediately the patient needs such as the reconsideration of current regimens of administration and the rational combination of drugs in use. In the medium-long term, due to new methodologies of medicinal-chemistry, the screening from natural products and the identification of new therapeutic targets, new lead compounds have great chance to advance through the drug development pipeline to clinic. Modern pharmaceutical formulation strategies and nanomedicines also merit a place in view of the benefits of a single dose of liposomal Amphotericin B (AmBisome®) against visceral leishmaniasis. BBB-targeted nanodevices could be suited for selective delivery of drugs against HAT encephalitic phase. Bioadhesive nanoparticles can be proposed to enhance the bioavailability of drugs after oral administration by reason of improving the drug solubility, and permeability across the intestinal epithelia.


Standard and biological treatment in large vessel vasculitis: guidelines and current approaches.
Giant cell arteritis and Takayasu arteritis are the two major forms of idiopathic large vessel vasculitis. High doses of glucocorticoids are effective in inducing remission in both conditions, but relapses and recurrences are common, requiring prolonged glucocorticoid treatment with the risk of the related adverse events. Areas covered: In this article, we will review the standard and biological treatment strategies in large vessel vasculitis, and we will focus on the current approaches to these diseases. Expert commentary: The results of treatment trials with conventional immunosuppressive agents such as methotrexate, azathioprine, mycophenolate mofetil, and cyclophosphamide have overall been disappointing. TNF-α blockers are ineffective in giant cell arteritis, while observational evidence and a phase 2 randomized trial support the use of tocilizumab in relapsing giant cell arteritis. Observational evidence strongly supports the use of anti-TNF-α agents and tocilizumab in Takayasu patients with relapsing disease. However biological agents are not curative, and relapses remain common.


Periadventitial Delivery of Simvastatin-Loaded Microparticles Attenuate Venous Neointimal Hyperplasia Associated With Arteriovenous Fistula.
Background Venous neointimal hyperplasia and venous stenosis (VS) formation can result in a decrease in arteriovenous fistula (AVF) patency in patients with end-stage renal disease. There are limited therapies that prevent VNH/VS. Systemic delivery of simvastatin has been shown to reduce VNH/VS but local delivery may help decrease the side effects associated with statin use. We determined if microparticles (MP) composed of cyclodextrins loaded with simvastatin (MP-SV) could reduce VS/VNH using a murine arteriovenous fistula model with chronic kidney disease. Methods and Results Male C57BL/6J mice underwent nephrectomy to induce chronic kidney disease. Four weeks later, an arteriovenous fistula was placed and animals were randomized to 3 groups: 20 μL of PBS or 20 μL of PBS with 16.6 mg/mL of either MP or MP-SV. Animals were euthanized 3 days later and the outflow veins were harvested for quantitative reverse transcriptase-polymerase chain reaction analysis and 28 days later for immunohistochemistical staining with morphometric analysis. Doppler ultrasound was performed weekly. Gene expression of vascular endothelial growth factor-A (Vegf-A), matrix metalloproteinase-9 (Mmp-9), transforming growth factor beta 1 (Tgf-β1), and monocyte chemoattractant protein-1 (Mcp-1) were significantly decreased in MP-SV treated vessels compared with controls. There was a significant decrease in the neointimal area, cell proliferation, inflammation, and fibrosis, with an increase in apoptosis and peak velocity in MP-SV treated outflow veins. MP-SV treated fibroblasts when exposed to hypoxic injury had decreased gene expression of Vegf-A and Mmp-9. Conclusions In experimental arteriovenous fistulas, periadventitial delivery of MP-SV decreased gene expression of Vegf-A, Mmp-9, Tgf-β1 and Mcp-1, VNH/VS, inflammation, and fibrosis.


Optic disc topographic analysis in diabetic patients.
The aim of our study was to evaluate the optic disc (OD) topographic parameters by using Heidelberg Retina Tomograph II (HRT II) in patients with type 2 diabetes mellitus (DM). The study group consisted of 78 patients with type 2 DM (patient group) and age-sex matched 50 healthy subjects (control group). All patients and controls underwent a detailed ophthalmological examination, automated perimetry, central corneal thickness (CCT) measurement and OD topography by using HRT II. Glycosylated haemoglobin (HbA1c) levels of the diabetic patients were also noted. Age, gender, intraocular pressure and CCT measurements were similar in patients with diabetes and control group. Mean retinal nerve fiber layer thickness (MRNFLT) and retinal nerve fiber layer cross-sectional area (RNFLcsA) parameters were found significantly lower in diabetic patients compared to those of the controls (p = 0.030, p = 0.038). In the patient group, MRNFLT value was found significantly lower in patients with DM duration ≥120 months (p = 0.020) and HbA1c level ≥7 % (p = 0.029). Rim volume, MRNFLT and RNFLcsA values were significantly lower in proliferative diabetic retinopathy group (p = 0.004, p = 0.003, p = 0.001 respectively) and laser treated patients (p = 0.003, p = 0.002, p = 0.004 respectively). In conclusion, poor metabolic control of diabetes, severe DR and received laser therapies cause RNFL damage. Heidelberg Retina Tomograph may help us to detect and follow-up the changes in optic disc and RNFL in diabetic patients.


Complementarity of solvent-free MALDI TOF and solid-state NMR spectroscopy in spectral analysis of polylactides.
We report systematic studies of solvent-free modification of matrix-assisted laser desorption/ionization time-of-flight (SF MALDI-TOF) mass spectrometry in analysis of synthetic polymers employing solid-state NMR spectroscopy as a supporting technique. In the present work oligomeric (M(n) = 4000 g mol(-1)) poly(L-lactide) (PLLA) was employed as a reference sample. The analyte was embedded into four matrixes commonly used in MALDI-TOF analysis of polymers: 1,8-dihydroxy-9-anthracenone (DT), 2,5-dihydroxybenzoic acid (DHB), 2-(4 hydroxyphenylazo)-benzoic acid (HABA), and trans-3-indoleacrylic acid (IAA). Solid-state NMR measurements clearly showed that the initial crystallinity of PLLA had no influence on quality of SF MALDI-TOF spectra since the crystalline structure of the analyte was not preserved during analyte/matrix grinding. Interestingly, the matrix remained crystalline during the sample's preparation. It was also found that, on the contrary to the dried droplet (DD) method, the SF approach leads to highly resolved mass spectra for a large variety of matrixes. Finally, problems of polymorphism and mechanochemical processes that can occur during the analyte/matrix grinding are briefly discussed.