B lymphocytes undergo TLR2-dependent apoptosis upon Shigella infection.
Antibody-mediated immunity to Shigella, the causative agent of bacillary dysentery, requires several episodes of infection to get primed and is short-lasting, suggesting that the B cell response is functionally impaired. We show that upon ex vivo infection of human colonic tissue, invasive S. flexneri interacts with and occasionally invades B lymphocytes. The induction of a type three secretion apparatus (T3SA)-dependent B cell death is observed in the human CL-01 B cell line in vitro, as well as in mouse B lymphocytes in vivo. In addition to cell death occurring in Shigella-invaded CL-01 B lymphocytes, we provide evidence that the T3SA needle tip protein IpaD can induce cell death in noninvaded cells. IpaD binds to and induces B cell apoptosis via TLR2, a signaling receptor thus far considered to result in activation of B lymphocytes. The presence of bacterial co-signals is required to sensitize B cells to apoptosis and to up-regulate tlr2, thus enhancing IpaD binding. Apoptotic B lymphocytes in contact with Shigella-IpaD are detected in rectal biopsies of infected individuals. This study therefore adds direct B lymphocyte targeting to the diversity of mechanisms used by Shigella to dampen the host immune response.


Phase II trials of dolastatin-10 in advanced pancreaticobiliary cancers.
Pancreaticobiliary malignancies respond poorly to conventional chemotherapy, and novel agents are needed. Dolatstatin-10 is a potent antimitotic pentapeptide isolated from the marine mollusk Dolabella auricularia that inhibits microtubule assembly. We conducted 2 parallel phase II trials of dolastatin-10 in patients with advanced hepatobiliary cancers and pancreatic adenocarcinoma. Eligible patients had histologically-confirmed metastatic pancreatic adenocarcinoma or metastatic, locally advanced or recurrent cancer of the liver, bile duct or gallbladder, and had received no prior chemotherapy for advanced disease. Dolastatin-10 400 microg/m(2) was administered intravenously by bolus every 21 days. Restaging CT scans were obtained every 2 cycles. Twenty-eight patients (16 hepatobiliary, including 7 hepatomas, 6 cholangiocarcinomas, 2 gallbladder carcinomas, and 12 pancreatic carcinomas) enrolled; 27 were evaluable for response. There were no objective responses. Grade 3/4 neutropenia occurred in 59% of patients and neutropenic fever in 18%. Median and 1-year survival were 5.0 months and 17% for the pancreatic cancer patients, and 3.0 months and 29% for the hepatobiliary patients. Median time to progression was 1.3 months for the pancreatic cancer patients and 1.6 months for the hepatobiliary patients. Dolastatin-10 is inactive against hepatobiliary and pancreatic carcinomas.


Whole exome sequencing identifies new causative mutations in Tunisian families with non-syndromic deafness.
Identification of the causative mutations in patients affected by autosomal recessive non syndromic deafness (DFNB forms), is demanding due to genetic heterogeneity. After the exclusion of GJB2 mutations and other mutations previously reported in Tunisian deaf patients, we performed whole exome sequencing in patients affected with severe to profound deafness, from four unrelated consanguineous Tunisian families. Four biallelic non previously reported mutations were identified in three different genes: a nonsense mutation, c.208C>T (p.R70X), in LRTOMT, a missense mutation, c.5417T>C (p.L1806P), in MYO15A and two splice site mutations, c.7395+3G>A, and c.2260+2T>A, in MYO15A and TMC1 respectively. We thereby provide evidence that whole exome sequencing is a powerful, cost-effective screening tool to identify mutations causing recessive deafness in consanguineous families.


Colony stimulating factors for prevention of myelosupressive therapy induced febrile neutropenia in children with acute lymphoblastic leukaemia.
Acute lymphoblastic leukaemia (ALL) is the most common cancer in childhood and febrile neutropenia is a potentially life-threatening side effect of its treatment. Current treatment consists of supportive care plus antibiotics. Clinical trials have attempted to evaluate the use of colony-stimulating factors (CSF) as additional therapy to prevent febrile neutropenia in children with ALL. The individual trials do not show whether there is significant benefit or not. Systematic review provides the most reliable assessment and the best recommendations for practice. To evaluate the safety and effectiveness of the addition of G-CSF or GM-CSF to myelosuppressive chemotherapy in children with ALL, in an effort to prevent the development of febrile neutropenia. Evaluation of number of febrile neutropenia episodes, length to neutrophil count recovery, incidence and length of hospitalisation, number of infectious disease episodes, incidence and length of treatment delays, side effects (flu-like syndrome, bone pain and allergic reaction), relapse and overall mortality (death). The search covered the Cochrane Central Register of Controlled Trials, MEDLINE, EMBASE, CANCERLIT, LILACS, and SciElo. We manually searched records of conference proceedings of ASCO and ASH from 1985 to 2003 as well as databases of ongoing trials. We consulted experts and scanned references from the relevant articles. We looked for randomised controlled trials (RCTs) comparing CSF with placebo or no treatment as primary or secondary prophylaxis to prevent febrile neutropenia in children with ALL. Two authors independently selected, critically appraised studies and extracted relevant data. The end points of interest were:* Primary end points: number of febrile neutropenia episodes and overall mortality (death) * Secondary end points: time to neutrophil count recovery, incidence and length of hospitalisation, number of infectious diseases episodes, incidence and length of treatment delays, side effects (flu-like syndrome, bone pain and allergic reaction) and relapse. We conducted a meta-analysis of these end points and expressed the results as Peto odds ratios. For continuous outcomes we calculated a weighted mean difference and a standardised mean difference. For count data, meta-analysis of the logarithms of the rate ratios using generic inverse variance was employed. We scanned more than 5500 citations and included six studies with a total of 332 participants in the analysis. There were insufficient data to assess the effect on survival. The use of CSF significantly reduced the number of episodes of febrile neutropenia episodes (Rate Ratio = 0.63; 95% confidence interval (CI) 0.46 to 0.85; p =0.003, with substantial heterogeneity), the length of hospitalisation (weighted mean difference (WMD) = -1.58; 95% CI -3.00 to -0.15; p = 0.03), and number of infectious diseases episodes (Rate Ratio=0.44; 95%CI 0.24 to 0.80; p=0.002). In spite of these results, CSF did not influence the length of episodes of neutropenia (WMD = -1.11; 95% CI -3.55 to 1.32; p = 0.4) or delays in chemotherapy courses (Rate Ratio=0.77; 95%CI 0.49 to 1,23; p=0.28) . Children with ALL treated with CSF benefit from shorter hospitalisation and fewer infections. However, there was no evidence for a shortened duration of neutropenia nor fewer treatment delays, and no useful information about survival. The role of CSF regarding febrile neutropenia episodes is still uncertain. Although current data shows statistical benefit for CSF use, substantial heterogeneity between included trials does not allow this conclusion.


Inhibition of the catalytic activity of alcohol dehydrogenase by nitric oxide is associated with S nitrosylation and the release of zinc.
Nitric oxide (NO) reacts with the sulfhydryl groups of proteins to form nitroso thiols. Alcohol dehydrogenase (ADH) plays an important role in the metabolism of ethanol. Chronic alcohol administration stimulates NO formation in the liver, and production of NO is increased in alcohol liver injury. The effect of exogenous and endogenous NO on rat or horse ADH activity was evaluated. Incubation of intact rat hepatocytes or cytosol isolated from hepatocytes with S-nitroso-N-acetylpenicillamine (SNAP), a nitric oxide donor, resulted in a decrease in ADH activity. Endogenous NO synthesis was induced in rat hepatocytes by incubation with a mixture of cytokines and endotoxin in the presence of L-arginine. As NO production in hepatocytes increased over a 24 h time period, a significant decrease in ADH activity was observed. This effect was blocked by the competitive inhibitor of NO synthesis, N omega-nitro-L-arginine methyl ester, indicating that ADH was also inactivated by endogenously generated NO. The decreased activity of ADH was not related to lowering of the ADH content as shown by Western blot analysis. To evaluate the mechanism of inhibition, purified ADH from equine liver was incubated with gaseous NO or NO released from NO donors such as the diethylamine/nitric oxide complex (DEA/NO) and SNAP. NO donors inactivated ADH in a dose- and time-dependent manner. Trapping of NO with hemoglobin resulted in protection of ADH against inactivation by NO. There was no effect by analogues of the NO donors which do not release NO. NAD afforded some protection against the NO inactivation of ADH. Measurements of thiol oxidation, S nitrosylation, and zinc release were used to assess the effect of NO on ADH activity. Thiol oxidation, S-nitroso thiol formation, and zinc release correlated with inactivation of ADH by NO, indicating that disruption of the zinc/thiolate active center due to S nitrosylation of ADH results in zinc release, followed by inactivation of the enzyme. Recovery experiments were performed by incubating the NO-treated enzyme with dithiothreitol (DTT) and/or Zn2+. The inhibitory effect by NO was reversible since, after the nitrosylated enzyme was reduced with DTT followed by incubation with ZnCl2 to allow reincorporation of Zn2+, ADH activity was increased from 20% of control values to 70%. These results suggest that cysteine residues contained within the zinc/thiolate active center may be primary sites of NO interaction with ADH. NO may modulate the metabolism of ethanol and influence metabolic actions of ethanol via interaction with ADH.


Treatment of experimental murine pancreatic peritoneal carcinomatosis with fibroblasts genetically modified to express IL12: a role for peritoneal innate immunity.
Peritoneal carcinomatosis from pancreatic cancer has a poor prognosis with a median survival of 3.1 months. This is mainly due to lack of effective treatment. Interleukin 12 (IL12) is a proinflammatory cytokine that has a potent antitumoral effect by stimulating innate and adoptive immunity. To examine the antitumoral effect and toxicity of intraperitoneal delivery of IL12 using an ex vivo gene therapy approach in a murine model of pancreatic peritoneal carcinomatosis. Peritoneal carcinomatosis was generated by direct intraperitoneal inoculation of the pancreatic cancer cell line Capan-1 in athymic mice. Syngenic fibroblasts were genetically modified in vitro to secrete IL12 using a polycistronic TFG murine IL12 retroviral vector coding for both p35 and p40 murine IL12 subunits. Ex vivo gene therapy involved injection of the genetically modified fibroblasts intraperitoneally twice a week for 4 weeks. Treatment of pre-established peritoneal carcinomatosis with fibroblasts genetically modified to express IL12 induced a marked inhibition of tumour growth as measured by comparison of the weights of the intraperitoneal tumour nodules in the treated and control animals (3.52 (SD 0.47) v 0.93 (SD 0.21) g, p<0.05) and improved survival. This effect was associated with infiltration of the peritoneal tumour nodules with macrophages. Peritoneal lavage confirmed enhancement of the innate peritoneal inflammatory activity, with an increased number of activated macrophages and natural killer cells. Moreover, macrophages harvested from animals with peritoneal carcinomatosis and treated with IL12-expressing fibroblasts expressed an activated proinflammatory antitumoral M1 phenotype that included strongly enhanced reactive oxygen species and nitric oxide production. There was no treatment-related toxicity. Multiple injections of genetically modified fibroblasts to express IL12 is an effective and well-tolerated treatment for experimental murine pancreatic peritoneal carcinomatosis via activated innate immunity and in particular activated M1 macrophages.


Modulation of spasticity: prolonged suppression of a spinal reflex by electrical stimulation.
Electrical subcutaneous nerve stimulation of radial, median, and saphenous nerves has been shown to produce prolonged analgesia. In a double blind study, such stimulation also suppressed clonus for 3 hours after stimulation ceased in subjects with spasticity. Since the effect is contralateral, each subject was his own control. Because stimulation of the nerve in the wrist suppressed ankle clonus, the mechanism mediating the effect must be centrifugal inhibition. These results suggest that subcutaneous nerve stimulation may also be a tool in the management of spasticity.


Bayesian analysis of parent-specific transmission ratio distortion in seven Spanish beef cattle breeds.
Transmission ratio distortion (TRD) is the departure from the expected Mendelian ratio in offspring, a poorly investigated biological phenomenon in livestock species. Given the current availability of specific parametric methods for the analysis of segregation data, this study focused on the screening of TRD in 602 402 single nucleotide polymorphisms covering all autosomal chromosomes in seven Spanish beef cattle breeds. On average, 0.13% (n = 786) and 0.01% (n = 29) of genetic markers evidenced sire- or dam-specific TRD respectively. There were no single nucleotide polymorphisms accounting for both sire- and dam-specific TRD at the same time, and only one marker (rs43147474) accounted for (sire-specific) TRD in all seven breeds. It must be noted that rs43147474 is located in the fourth intronic region of the GTP-binding protein 10 gene, and this locus has been previously linked to the maintenance of mitochondria and nucleolar architectures. Alternatively, other candidate genes surround this hot-spot for sire-specific TRD in the cattle genome, and they are related to embryonic and postnatal lethality as well as prostate cancer, among others. This research characterized the distribution of TRD in the bovine genome, highlighting heterogeneous results when comparing across breeds.


Transmembrane protein 198 promotes LRP6 phosphorylation and Wnt signaling activation.
Wnt/β-catenin signaling is fundamental in embryogenesis and tissue homeostasis in metazoans. Upon Wnt stimulation, cognate coreceptors LRP5 and LRP6 ([LRP5/6] low-density lipoprotein receptor-related proteins 5 and 6) are activated via phosphorylation at key residues. Although several kinases have been implicated, the LRP5/6 activation mechanism remains unclear. Here, we report that transmembrane protein 198 (TMEM198), a previously uncharacterized seven-transmembrane protein, is able to specifically activate LRP6 in transducing Wnt signaling. TMEM198 associates with LRP6 and recruits casein kinase family proteins, via the cytoplasmic domain, to phosphorylate key residues important for LRP6 activation. In mammalian cells, TMEM198 is required for Wnt signaling and casein kinase 1-induced LRP6 phosphorylation. During Xenopus embryogenesis, maternal and zygotic tmem198 mRNAs are widely distributed in the ectoderm and mesoderm. TMEM198 is required for Wnt-mediated neural crest formation, antero-posterior patterning, and particularly engrailed-2 expression in Xenopus embryos. Thus, our results identified TMEM198 as a membrane scaffold protein that promotes LRP6 phosphorylation and Wnt signaling activation.


Measuring asthma control. Clinic questionnaire or daily diary?
Daily symptom, peak expiratory flow rate (PEFR), and medication diaries are often used in clinical trials of treatments for asthma on the assumption that they provide a better estimate of clinical status than does a questionnaire completed in the clinic. We conducted a study with the aim of comparing the measurement properties of the clinic-completed Asthma Control Questionnaire with those of the Asthma Control Diary. The diary is composed of questions and response options almost identical to those of the questionnaire, but uses PEFR instead of FEV(1) as the measure of airway caliber. In an observational study, 50 adults with symptomatic asthma attended a McMaster University asthma clinic at 0, 1, 5, and 9 wk to complete the Asthma Control Questionnaire and other measures of asthma status. For 1 wk before each follow-up visit, patients completed the Asthma Control Diary every morning and evening. Concordance between the questionnaire and diary was high (intraclass correlation coefficient [ICC] = 0.87). Both reliability (ICC: questionnaire = 0.90; diary = 0.86) and responsiveness (responsiveness index: questionnaire = 1.06; diary = 0.90; p = 0.005) were better with the questionnaire than with the diary. Correlations between the two instruments and other measures of clinical asthma status were similar and close to a priori predictions. Both the Asthma Control Questionnaire and the Asthma Control Diary are valid instruments for measuring asthma control, but the questionnaire has slightly better discriminative and evaluative measurement properties than does the diary.


[Acute appendicitis caused by Balantidium coli].
A patient who was surgically treated for acute appendicitis is presented. In the sections of cecal appendix many Balantidium coli trophozoites were found. The history, characteristics, habitat, location, biological aspects and reproduction of this parasite are commented.


Dynamics of transparent exopolymer particle production and aggregation during viral infection of the coccolithophore, Emiliania huxleyi.
Emiliania huxleyi produces calcium carbonate (CaCO3 ) coccoliths and transparent exopolymer particles (TEP), sticky, acidic carbohydrates that facilitate aggregation. E. huxleyi's extensive oceanic blooms are often terminated by coccolithoviruses (EhVs) with the transport of cellular debris and associated particulate organic carbon (POC) to depth being facilitated by TEP-bound 'marine snow' aggregates. The dynamics of TEP production and particle aggregation in response to EhV infection are poorly understood. Using flow cytometry, spectrophotometry and FlowCam visualization of alcian blue (AB)-stained aggregates, we assessed TEP production and the size spectrum of aggregates for E. huxleyi possessing different degrees of calcification and cellular CaCO3 :POC mass ratios, when challenged with two EhVs (EhV207 and EhV99B1). FlowCam imaging also qualitatively assessed the relative amount of AB-stainable TEP (i.e., blue:red ratio of each particle). We show significant increases in TEP during early phase EhV207-infection (∼ 24 h) of calcifying strains and a shift towards large aggregates following EhV99B1-infection. We also observed the formation of large aggregates with low blue:red ratios, suggesting that other exopolymer substances contribute towards aggregation. Our findings show the potential for virus infection and the associated response of their hosts to impact carbon flux dynamics and provide incentive to explore these dynamics in natural populations.


Branching out: mechanisms of dendritic arborization.
Type-specific dendrite morphology is a hallmark of the neuron and has important functional implications in determining what signals a neuron receives and how these signals are integrated. During the past two decades, studies on dendritic arborization neurons in Drosophila melanogaster have started to identify mechanisms of dendrite morphogenesis that may have broad applicability to vertebrate species. Transcription factors, receptor-ligand interactions, various signalling pathways, local translational machinery, cytoskeletal elements, Golgi outposts and endosomes have been identified as contributors to the organization of dendrites of individual neurons and the placement of these dendrites in the neuronal circuitry. Further insight into these mechanisms will improve our understanding of how the nervous system functions and might help to identify the underlying causes of some neurological and neurodevelopmental disorders.


Experimental evidence of a buoyant mass difference between bovine spermatozoa bearing X- and Y-chromosomes using a micromechanical resonator.
Flow cytometry is to date the only commercially viable technique for sex preselection of mammalian spermatozoa, measuring the different DNA content in X- and Y-chromosome bearing spermatozoa. Here we present experimental evidence of a measurable difference between bovine spermatozoa bearing X- and Y-chromosomes based on their buoyant mass. Single cells of two populations of flow-cytometrically sorted spermatozoa were analyzed by means of a micromechanical resonator, consisting of a suspended doubly-clamped microcapillary. Spermatozoa buoyant mass is related to the transitory variation in vibration phase lag, caused by the passage through the sensitive area of a single sperm cell suspended in a fluid. Data analysis shows two well-separated distributions and provides evidence of the sensor capabilities to detect the buoyant mass of single cells with such accuracy to distinguish X- and Y-chromosome bearing spermatozoa. These preliminary results suggest the possibility to develop an intriguing technique alternative to flow cytometry in the field of sperm sorting.


Anthocyanin-enriched extract from Ribes nigrum inhibits triglyceride and cholesterol accumulation in adipocytes.
Aim: Obesity is a chronic pathology of epidemic proportions. Mature adipocytes from a 3T3-L1 cell line were used as in vitro obesity model to test different bioactive compounds. We aim to evaluate cassis (Ribes nigrum) extract antioxidant activity and its antiadipogenic effect on mature adipocytes. Results: We produced an extract by using enzyme that combines cellulase and pectinase; we obtained high yield of the bioactive compound anthocyanin. Extract showed high antioxidant capacity. We conducted in vitro assays by adding the extract to adipocytes culture medium. Extract reduced intracellular levels of triglyceride by 62% and cholesterol by 32%. Conclusion: Enzymatic extract's high antioxidant activity was likely attributable to its high concentration of anthocyanin. This extract inhibits lipid accumulation in adipocytes.


Localization of a candidate surfactant convertase to type II cells, macrophages, and surfactant subfractions.
Pulmonary surfactant exists in the alveolus in several distinct subtypes that differ in their morphology, composition, and surface activity. Experiments by others have implicated a serine hydrolase in the production of the inactive small vesicular subtype of surfactant (N. J. Gross and R. M. Schultz. Biochim. Biophys. Acta 1044: 222-230, 1990). Our laboratory recently identified this enzyme in the rat as the serine carboxylesterase ES-2 [F. Barr, H. Clark, and S. Hawgood. Am. J. Physiol. 274 (Lung Cell. Mol. Physiol. 18): L404-L410, 1998]. In the present study, we determined the cellular sites of expression of ES-2 in rat lung using a digoxygenin-labeled ES-2 riboprobe. ES-2 mRNA was localized to type II cells and alveolar macrophages but not to Clara cells. Using a specific ES-2 antibody, we determined the protein distribution of ES-2 in the lung by immunohistochemistry, and it was found to be consistent with the sites of mRNA expression. Most of the ES-2 in rat bronchoalveolar lavage is in the surfactant-depleted supernatant, but ES-2 was also consistently localized to the small vesicular surfactant subfraction presumed to form as a consequence of conversion activity. These results are consistent with a role for endogenous lung ES-2 in surfactant metabolism.


Impulse Data Models for the Inverse Problem of Electrocardiography.
To develop, train and test neural networks for predicting heart surface potentials (HSPs) from body surface potentials (BSPs). The method re-frames traditional inverse problems of electrocardiography into regression problems, constraining the solution space by decomposing signals with multidimensional Gaussian impulse basis functions. Impulse HSPs were generated with single Gaussian basis functions at discrete heart surface locations and projected to corresponding BSPs using a volume conductor torso model. Both BSP (inputs) and HSP (outputs) were mapped to regular 2D surface meshes and used to train a neural network. Predictive capabilities of the network were tested with unseen synthetic and experimental data. A dense full connected single hidden layer neural network was trained to map body surface impulses to heart surface Gaussian basis functions for reconstructing HSP. Synthetic pulses moving across the heart surface were predicted from the neural network with root mean squared error of 9.1±1.4%. Predicted signals were robust to noise up to 20 dB and errors due to displacement and rotation of the heart within the torso were bounded and predictable. A shift of the heart 40 mm toward the spine resulted in a 4% increase in signal feature localization error. The set of training impulse function data could be reduced, and prediction error remained bounded. Recorded HSPs from in-vitro pig hearts were reliably decomposed using space-time Gaussian basis functions. Activation times calculated from predicted HSPs for left-ventricular pacing had a mean absolute error of 10.4±11.4 ms. Other pacing scenarios were analyzed with similar success. Impulses from Gaussian basis functions are potentially an effective and robust way to train simple neural network data models for reconstructing HSPs from decomposed BSPs. The HSPs predicted by the neural network can be used to generate activation maps that non-invasively identify features of cardiac electrical dysfunction and can guide subsequent treatment options.


Pharmacokinetic Profile of Gilteritinib: A Novel FLT-3 Tyrosine Kinase Inhibitor.
Gilteritinib is a novel, highly selective tyrosine kinase inhibitor approved in the USA, Canada, Europe, Brazil, Korea, and Japan for the treatment of FLT3 mutation-positive acute myeloid leukemia. This article describes the clinical pharmacokinetic profile of gilteritinib. The pharmacokinetic profile of gilteritinib was assessed from five clinical studies. Dose-proportional pharmacokinetics was observed following once-daily gilteritinib administration (dose range 20-450 mg). Median maximum concentration was reached 2-6 h following single and repeat dosing of gilteritinib; mean elimination half-life was 113 h. Elimination was primarily via feces. Exposure to gilteritinib was comparable under fasted and fed conditions. Gilteritinib is primarily metabolized via cytochrome P450 (CYP) 3A4; coadministration of gilteritinib with itraconazole (a strong P-glycoprotein inhibitor and CYP3A4 inhibitor) or rifampicin (a strong P-glycoprotein inducer and CYP3A inducer) significantly affected the gilteritinib pharmacokinetic profile. No clinically relevant interactions were observed when gilteritinib was coadministered with midazolam (a CYP3A4 substrate) or cephalexin (a multidrug and toxin extrusion 1 substrate). Unbound gilteritinib exposure was similar between subjects with hepatic impairment and normal hepatic function. Gilteritinib exhibits a dose-proportional pharmacokinetic profile in healthy subjects and in patients with relapsed/refractory acute myeloid leukemia. Gilteritinib exposure is not significantly affected by food. Moderate-to-strong CYP3A inhibitors demonstrated a significant effect on gilteritinib exposure. Coadministration of gilteritinib with CYP3A4 or multidrug and toxin extrusion 1 substrates did not impact substrate concentrations. Unbound gilteritinib was comparable between subjects with hepatic impairment and normal hepatic function; dose adjustment is not warranted for patients with hepatic impairment. NCT02014558, NCT02456883, NCT02571816.


FACS-optimized mutants of the green fluorescent protein (GFP).
We have constructed a library in Escherichia coli of mutant gfp genes (encoding green fluorescent protein, GFP) expressed from a tightly regulated inducible promoter. We introduced random amino acid (aa) substitutions in the twenty aa flanking the chromophore Ser-Tyr-Gly sequence at aa 65-67. We then used fluorescence-activated cell sorting (FACS) to select variants of GFP that fluoresce between 20-and 35-fold more intensely than wild type (wt), when excited at 488 nm. Sequence analysis reveals three classes of aa substitutions in GFP. All three classes of mutant proteins have highly shifted excitation maxima. In addition, when produced in E. coli, the folding of the mutant proteins is more efficient than folding of wt GFP. These two properties contribute to a greatly increased (100-fold) fluorescence intensity, making the mutants useful for a number of applications.


Acoustic radiation force impulse imaging: in vivo demonstration of clinical feasibility.
The clinical viability of a method of acoustic remote palpation, capable of imaging local variations in the mechanical properties of soft tissue using acoustic radiation force impulse (ARFI) imaging, is investigated in vivo. In this method, focused ultrasound (US) is used to apply localized radiation force to small volumes of tissue (2 mm(3)) for short durations (less than 1 ms) and the resulting tissue displacements are mapped using ultrasonic correlation-based methods. The tissue displacements are inversely proportional to the stiffness of the tissue and, thus, a stiffer region of tissue exhibits smaller displacements than a more compliant region. Due to the short duration of the force application, this method provides information about the mechanical impulse response of the tissue, which reflects variations in tissue viscoelastic characteristics. In this paper, experimental results are presented demonstrating that displacements on the order of 10 microm can be generated and detected in soft tissues in vivo using a single transducer on a modified diagnostic US scanner. Differences in the magnitude of displacement and the transient response of tissue are correlated with tissue structures in matched B-mode images. The results comprise the first in vivo ARFI images, and support the clinical feasibility of a radiation force-based remote palpation imaging system.


False lumen pressure estimation in type B aortic dissection using 4D flow cardiovascular magnetic resonance: comparisons with aortic growth.
Chronic type B aortic dissection (TBAD) is associated with poor long-term outcome, and accurate risk stratification tools remain lacking. Pressurization of the false lumen (FL) has been recognized as central in promoting aortic growth. Several surrogate imaging-based metrics have been proposed to assess FL hemodynamics; however, their relationship to enlarging aortic dimensions remains unclear. We investigated the association between aortic growth and three cardiovascular magnetic resonance (CMR)-derived metrics of FL pressurization: false lumen ejection fraction (FLEF), maximum systolic deceleration rate (MSDR), and FL relative pressure (FL ΔPmax). CMR/CMR angiography was performed in 12 patients with chronic dissection of the descending thoracoabdominal aorta, including contrast-enhanced CMR angiography and time-resolved three-dimensional phase-contrast CMR (4D Flow CMR). Aortic growth rate was calculated as the change in maximal aortic diameter between baseline and follow-up imaging studies over the time interval, with patients categorized as having either 'stable' (< 3 mm/year) or 'enlarging' (≥ 3 mm/year) growth. Three metrics relating to FL pressurization were defined as: (1) FLEF: the ratio between retrograde and antegrade flow at the TBAD entry tear, (2) MSDR: the absolute difference between maximum and minimum systolic acceleration in the proximal FL, and (3) FL ΔPmax: the difference in absolute pressure between aortic root and distal FL. FLEF was higher in enlarging TBAD (49.0 ± 17.9% vs. 10.0 ± 11.9%, p = 0.002), whereas FL ΔPmax was lower (32.2 ± 10.8 vs. 57.2 ± 12.5 mmHg/m, p = 0.017). MSDR and conventional anatomic variables did not differ significantly between groups. FLEF showed positive (r = 0.78, p = 0.003) correlation with aortic growth rate whereas FL ΔPmax showed negative correlation (r = - 0.64, p = 0.026). FLEF and FL ΔPmax remained as independent predictors of aortic growth rate after adjusting for baseline aortic diameter. Comparative analysis of three 4D flow CMR metrics of TBAD FL pressurization demonstrated that those that focusing on retrograde flow (FLEF) and relative pressure (FL ΔPmax) independently correlated with growth and differentiated patients with enlarging and stable descending aortic dissections. These results emphasize the highly variable nature of aortic hemodynamics in TBAD patients, and suggest that 4D Flow CMR derived metrics of FL pressurization may be useful to separate patients at highest and lowest risk for progressive aortic growth and complications.


Vector length as a proxy for the adequacy of ultrafiltration in hemodialysis.
Evaluation of dialysis adequacy has focused on parameters of solute (principally urea) clearance. Relatively little attention has been paid to the adequacy of ultrafiltration. At a given phase angle, the bioimpedance vector length reflects the degree of tissue hydration, as the vector lengthens with ultrafiltration. We determined the relative risk of death associated with different bioimpedance vector lengths in a 3009 patient hemodialysis cohort using proportional hazards regression. The mean phase angle was 4.8 degrees, and the mean vector length 300 +/- 70 ohm/m (range 140 to 630 ohm/m). Vector length was much longer in women than men (mean 340 vs. 270 ohm/m) and significantly longer in African Americans and patients without diabetes. Adjusted for the effects of age, gender, race, diabetes, vintage, weight, albumin, prealbumin, creatinine, hemoglobin, ferritin, and dialysis dose, the relative risk (RR) of death was 0.75 (95% CI 0.57 to 0.88) per 100 ohm/m decrease in vector length. The effect of vector length on RR was somewhat more pronounced among men (vector length x gender interaction, P= 0.07). Considering vector length of 300 to 350 ohm/m as the referent category, the RRs of death were 1.54 (95% CI 1.08 to 2.21) and 2.83 (95% CI 1.55 to 5.14) for patients with vector length 200 to 250 and <200 ohm/m, respectively. Shorter predialysis bioimpedance vectors, indicating greater soft tissue hydration, were associated with diminished survival in hemodialysis patients. These findings validate clinical observations linking longevity to maintenance of dry body weight.


The advantages of deep ocean water for the development of functional fermentation food.
Deep ocean water (DOW) is obtained from 600 m below the sea surface. In recent years, DOW has been applied in the development of fermentation biotechnologies and functional foods. DOW is rich in trace minerals, comprises multiple physiological and health functions, and is able to promote microbe growth; therefore, the application of DOW directly benefits the development of the fermentation industry and functional foods. This study integrated the current health functions and applications of DOW with the latest results from studies related to fermentation biotechnology. Subsequently, the influence of applying DOW in fermented functional food development and the effects in health function improvements were summarized. According to the previous studies, the main reasons for the increased effect of fermented functional foods through the application of DOW are increased generation of functional metabolite contents in the microbes, intrinsic health functions of DOW, and the microbial use of mechanisms of converting the absorbed inorganic ions into highly bioavailable organic ions for the human body. These combined advantages not only enhance the health functions of fermentation products but also provide fermentation products with the intrinsic health functions of DOW.


Effect of saponin treatment on the sarcoplasmic reticulum of rat, cane toad and crustacean (yabby) skeletal muscle.
1. Mechanically skinned fibres from skeletal muscles of the rat, toad and yabby were used to investigate the effect of saponin treatment on sarcoplasmic reticulum (SR) Ca2+ loading properties. The SR was loaded submaximally under control conditions before and after treatment with saponin and SR Ca2+ was released with caffeine. 2. Treatment with 10 micrograms ml-1 saponin greatly reduced the SR Ca2+ loading ability of skinned fibres from the extensor digitorum longus muscle of the rat with a rate constant of 0.24 min-1. Saponin concentrations up to 150 micrograms ml-1 and increased exposure time up to 30 min did not further reduce the SR Ca2+ loading ability of the SR, which indicates that the inhibitory action of 10-150 micrograms ml-1 saponin is not dose dependent. The effect of saponin was also not dependent on the state of polarization of the transverse-tubular system. 3. Treatment with saponin at concentrations up to 100 micrograms ml-1 for 30 min did not affect the Ca2+ loading ability of SR in skinned skeletal muscle fibres from the twitch portion of the toad iliofibularis muscle but SR Ca2+ loading ability decreased markedly with a time constant of 0.22 min-1 in the presence of 150 micrograms ml-1 saponin. 4. The saponin dependent increase in permeability could be reversed in both rat and toad fibres by short treatment with 6 microM Ruthenium Red, a potent SR Ca2+ channel blocker, suggesting that saponin does affect the SR Ca2+ channel properties in mammalian and anuran skeletal muscle. 5. Treatment of skinned fibres of long sarcomere length (> 6 microns) from the claw muscle of the yabby (a freshwater decapod crustacean) with 10 micrograms ml-1 saponin for 30 min abolished the ability of the SR to load Ca2+, indicating that saponin affects differently the SR from skeletal muscles of mammals, anurans and crustaceans. 6. It is concluded that at relatively low concentrations, saponin causes inhibition of the skeletal SR Ca2+ loading ability in a species dependent manner, probably by increasing the Ca2+ loss through SR Ca2+ release channels.


Regulatory T cells selectively control CD8+ T cell effector pool size via IL-2 restriction.
Regulatory T cells (Treg) are key players in maintaining immune homeostasis but have also been shown to regulate immune responses against infectious pathogens. Therefore, Treg are a promising target for modulating immune responses to vaccines to improve their efficacy. Using a viral vector system, we found that Treg act on the developing immune response early postinfection by reducing the extent of dendritic cell costimulatory molecule expression. Due to this change and the lower IL-2 production that results, a substantial fraction of CD8(+) effector T cells lose CD25 expression several days after activation. Surprisingly, such Treg-dependent limitations in IL-2 signaling by Ag-activated CD8(+) T cells prevent effector differentiation without interfering with memory cell formation. In this way, Treg fine-tune the numbers of effector T cells generated while preserving the capacity for a rapid recall response upon pathogen re-exposure. This selective effect of Treg on a subpopulation of CD8(+) T cells indicates that although manipulation of the Treg compartment might not be optimal for prophylactic vaccinations, it can be potentially exploited to optimize vaccine efficacy for therapeutic interventions.


Elucidation of the Aggregation Pathways of Helix-Turn-Helix Peptides: Stabilization at the Turn Region Is Critical for Fibril Formation.
Aggregation of proteins to fiberlike aggregates often involves a transformation of native monomers to β-sheet-rich oligomers. This general observation underestimates the importance of α-helical segments in the aggregation cascade. Here, using a combination of experimental techniques and accelerated molecular dynamics simulations, we investigate the aggregation of a 43-residue, apolipoprotein A-I mimetic peptide and its E21Q and D26N mutants. Our study indicates a strong propensity of helical segments not to adopt cross-β-fibrils. The helix-turn-helix monomeric conformation of the peptides is preserved in the mature fibrils. Furthermore, we reveal opposite effects of mutations on and near the turn region in the self-assembly of these peptides. We show that the E21-R24 salt bridge is a major contributor to helix-turn-helix folding, subsequently leading to abundant fibril formation. On the other hand, the K19-D26 interaction is not required to fold the native helix-turn-helix peptide. However, removal of the charged D26 residue decreases the stability of the helix-turn-helix monomer and consequently reduces the level of aggregation. Finally, we provide a more refined assembly model for the helix-turn-helix peptides from apolipoprotein A-I based on the parallel stacking of helix-turn-helix dimers.


caOmicsV: an R package for visualizing multidimensional cancer genomic data.
Translational genomics research in cancers, e.g., International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), has generated large multidimensional datasets from high-throughput technologies. Data analysis at multidimensional level will greatly benefit clinical applications of genomic information in diagnosis, prognosis and therapeutics of cancers. To help, tools to effectively visualize integrated multidimensional data are important for understanding and describing the relationship between genomic variations and cancers. We implemented the R package, caOmicsV, to provide methods under R environment to visualize multidimensional cancer genomic data in two layouts: matrix layout and combined biological network and circular layout. Both layouts support to display sample information, gene expression (e.g., RNA and miRNA), DNA methylation, DNA copy number variations, and summarized data. A set of supplemental functions are included in the caOmicsV package to help users in generation of plot data sets from multiple genomic datasets with given gene names and sample names. Default plot methods for both layouts for easy use are also implemented. caOmicsV package provides an easy and flexible way to visualize integrated multidimensional cancer genomic data under R environment.


Modulating SIRT1, Nrf2, and NF-κB signaling pathways by bergenin ameliorates the cadmium-induced nephrotoxicity in rats.
In light of the current industrial evolution, exposure to cadmium has become a significant public health concern. Cadmium accumulates in the renal tubular cells and causes nephrotoxicity largely through disruption of the redox homeostasis, induction of inflammation, and suppression of the histone deacetylase SIRT1 expression. The current work aimed at exploring the protective capability of bergenin, a naturally-occurring methyl gallic acid derivative, against the cadmium-evoked nephrotoxicity. Male Wistar rats were treated either with cadmium alone or with cadmium and bergenin for a 7-day experimental period followed by collection of kidney and blood specimens that were subjected to biochemical, molecular, and histological investigations. The results revealed the ability of bergenin to improve the renal functions in the cadmium-intoxicated rats as evidenced by increased glomerular filtration rate, and decreased serum creatinine and blood urea nitrogen. Equally important, bergenin reduced the renal tissue injury and enhanced its redox homeostasis as indicated by decreased protein expression of the kidney injury marker KIM-1, reduced lipid peroxidation, and improved antioxidant potential and histopathological picture of the renal tissues. Mechanistically, bergenin reduced the renal tissue cadmium content, markedly up-regulated protein expression of SIRT1 that regulates inflammation and the redox status of the renal tissues. Additionally, it improved the expression of the major antioxidant transcription factor Nrf2 and its responsive gene products heoxygenase-1 and NAD(P)H quinone dehydrogenase 1 in the cadmium-intoxicated rats. In the same context, bergenin down-regulated the acetylation and the nuclear translocation of the inflammatory transcription factor NF-κB and reduced levels of its responsive gene products TNF-α and IL-1β, as well as the activity of the inflammatory cell infiltration biomarker myeloperoxidase. Collectively, the current study underscores the ameliorating activity of bergenin against the cadmium-evoked nephrotoxicity and highlights modulation of SIRT1, Nrf2, and NF-κB signaling as potential underlining molecular mechanisms.


Broad spectrum diffusion model for room acoustics ray-tracing algorithms
Geometrical acoustics methods have already been transformed to account for diffusely reflecting boundaries. In randomized ray-tracing algorithms, the sound rays are either specularly reflected or scattered, according to the value of a random number which is compared with the diffusion factor. However, this method becomes inefficient if this factor depends on frequency, since the process must then be repeated for each frequency band. A method is proposed in this paper to compute all frequency components simultaneously in a single pass. The diffuse reflection model is based on the definition of a new concept: the "splitting coefficient," which can differ from the diffusion factor. First, the randomized ray-tracing method is described and the diffusion model is applied to a single diffusing surface in free field. It is shown that the results computed by the program are in accordance with theoretical results. Then, the method is shown to work properly when it is applied to more realistic enclosures: This is proved in theory and illustrated by some examples. A particular problem is the increase of the statistical error which has been solved by an appropriate control of the splitting coefficient.


Lung ultrasound patterns in paediatric pneumonia in Mozambique and Pakistan.
Improved pneumonia diagnostics are needed, particularly in resource-constrained settings. Lung ultrasound (LUS) is a promising point-of-care imaging technology for diagnosing pneumonia. The objective was to explore LUS patterns associated with paediatric pneumonia. We conducted a prospective, observational study among children aged 2 to 23 months with World Health Organization Integrated Management of Childhood Illness chest-indrawing pneumonia and among children without fast breathing, chest indrawing or fever (no pneumonia cohort) at two district hospitals in Mozambique and Pakistan. We assessed LUS and chest radiograph (CXR) examinations, and viral and bacterial nasopharyngeal carriage, and performed a secondary analysis of LUS patterns. LUS demonstrated a range of distinctive patterns that differed between children with and without pneumonia and between children in Mozambique versus Pakistan. The presence of LUS consolidation or interstitial patterns was more common in children with chest-indrawing pneumonia than in those without pneumonia. Consolidations were also more common among those with only bacterial but no viral carriage detected (50.0%) than among those with both (13.0%) and those with only virus detected (8.3%; p=0.03). LUS showed high interrater reliability among expert LUS interpreters for overall determination of pneumonia (κ=0.915), consolidation (κ=0.915) and interstitial patterns (κ=0.901), but interrater reliability between LUS and CXR for detecting consolidations was poor (κ=0.159, Pakistan) to fair (κ=0.453, Mozambique). Pattern recognition was discordant between LUS and CXR imaging modalities. Further research is needed to define and standardise LUS patterns associated with paediatric pneumonia and to evaluate the potential value of LUS as a reference standard.


NADPH oxidase and nitric oxide synthase-dependent superoxide production is increased in proliferative lupus nephritis.
Lupus nephritis (LN) is an immune complex-mediated glomerulonephritis. Proliferative LN (PLN, ISN/RPS classes III and IV)) often leads to renal injury or failure despite traditional induction and maintenance therapy. Successful targeted therapeutic development requires insight into mediators of inflammation in PLN. Superoxide (SO) and its metabolites are mediators of the innate immune response through their ability to mediate reduction-oxidation signaling. Endothelial nitric oxide synthase (eNOS) modulates inflammatory responses in endothelial cells. We hypothesized that markers of SO production would be increased in active PLN and that SO production would be dependent on the activity of select enzymes in the renal cortex. Patients with systemic lupus erythematosus were enrolled at the time of renal biopsy for active LN of all classes. Serum collected at baseline was analyzed by HPLC with electrochemical detection for markers of SO production (durable modifications of serum protein Tyr ultimately requiring SO as a substrate). Renal cortex from MRL/MpJ-FAS(lpr) (MRL/lpr) mice with and without functional eNOS was analyzed during active disease for superoxide (SO) production with and without inhibitors of SO-producing enzymes. Serum protein modifications indicative of total SO production were significantly higher in patients with PLN. These markers were increased in association with more active, inflammatory PLN. Mice lacking functional eNOS had 80% higher levels of renal cortical SO during active disease, and inhibitors of nitric oxide synthase and NADPH oxidase reduced these levels by 60% and 77%, respectively. These studies demonstrate that SO production is unique to active PLN in a NOS and NADPH oxidase-dependent fashion. These findings suggest the emulating or augmenting eNOS activity or inhibiting NADPH oxidase SO production may be targets of therapy in patients with PLN. The markers of SO production used in this study could rationally be used to select SO-modulating therapies and serve as pharmacodynamic indicators for dose titration.


Marine brevetoxin induces IgE-independent mast cell activation.
Brevetoxins (PbTx) are sodium channel neurotoxins produced by the marine dinoflagellate Karenia brevis during red tide blooms. Inhalation of PbTx in normal individuals and individuals with pre-existing airways disease results in adverse airway symptoms including bronchoconstriction. In animal models of allergic inflammation, inhalation of PbTx results in a histamine H₁-mediated bronchoconstriction suggestive of mast cell activation. How mast cells would respond directly to PbTx is unknown. We thus explored the activation of mouse bone marrow-derived mast cells (BMMCs) following exposure to purified PbTx-2. Following in vitro exposure to PbTx-2, we examined cellular viability, mast cell degranulation (β-hexosaminidase release), intracellular Ca²+ and Na+ flux, and the production of inflammatory mediators (IL-6). PbTx-2 induced significant cellular toxicity within 24 h as measured by LDH release and Annexin-V staining. However, within 1 h of exposure, PbTx-2 induced BMMC degranulation and an increase in IL-6 mRNA expression independent of the high-affinity IgE receptor (FcεRI) stimulation. Activation of BMMCs by PbTx-2 was associated with altered intracellular Ca²+ and Na+ levels. Brevenal, a naturally produced compound that antagonizes the activity of PbTx, prevented changes in intracellular Na+ levels but did not alter activation of BMMCs by PbTx-2. These findings demonstrate that PbTx-2 activates mast cells independent of FcεRI providing insight into critical events in the pathogenesis and a potential therapeutic target in brevetoxin-induced airway symptoms.


Mediator of transcriptional regulation.
A multi-protein complex, termed mediator, has been isolated from yeast, based on its requirement for transcriptional activation in a system reconstituted from pure RNA polymerase II and general transcription factors. Mediator polypeptides include the products of many genes previously recovered from screens for mutations affecting transcription. This connection between biochemical and genetic studies reveals that mediator is important for both activation and repression of transcription, and that mediator plays a role in transcriptional regulation in vivo as well as in vitro. Mediator binds the carboxy-terminal domain of RNA polymerase II, forming a polymerase holoenzyme, whose possible association with additional proteins is a subject of some controversy.


Effect of light curing protocol on degree of conversion of composites.
To evaluate the degree of conversion (DC) of two light-cured composites with different protocols of light curing. One hundred and ninety two specimens (n = 8) were prepared (5 mm × 2 mm) according to experimental groups: two composite resins (Filtek Supreme and four seasons); three light curing protocols [20 s with the tip of the light curing unit (LCU) device touching composite surface (C); 20 s with the tip of the LCU at 8 mm distant from composite surface (D); and tip of the LCU at 8 mm distant from composite surface and polymerization time required to obtain a radiant exposure of 16 J/cm(2) (DS)]. Four LCUs (Bluephase 16i, Ultralume LED 5, XL 3000 and Optilux 501C) were used. DC of the bottom and top surface of specimens were measured using a FTIR spectrometer. Data were statistically analyzed by 3-way split splot ANOVA and Tukey's test (alpha = 0.05). The results showed that DC of the top surface was higher than the bottom at all experimental conditions (p < 0.0001). Overall, the curing at 8 mm of distance did not affect conversion rate on the top surface (p > 0.05), but bottom surfaces showed DC reduction (p < 0.05). The highest monomer conversion values were observed for C and DS situations. The distance between the LCU and material surface and radiant exposure can affect the DC. Polymerization at distance should be performed with curing units with higher light power and/or extended exposure time.


A novel use of reticulated hyaluronic acid (Healaflow) for hypotony eyes in patients with uveitis.
Persistent ocular hypotony is a complex and ongoing challenge faced in ophthalmology. It can result in early ocular phthisis and associated visual decline, pain and deformity. We present the first case series, in which repeated intracameral injections of highly reticulated hyaluronic acid (Healaflow) have successfully prevented the complications of ocular hypotony in the long term. We believe it is a viable management option that can bring about a significant improvement to the quality of life in this subgroup of patients while avoiding frequent intervention.


Characterisation of cell cycle arrest and terminal differentiation in a maximally proliferative human epithelial tissue: Lessons from the human hair follicle matrix.
Human hair follicle (HF) growth and hair shaft formation require terminal differentiation-associated cell cycle arrest of highly proliferative matrix keratinocytes. However, the regulation of this complex event remains unknown. CIP/KIP family member proteins (p21CIP1, p27KIP1 and p57KIP2) regulate cell cycle progression/arrest, endoreplication, differentiation and apoptosis. Since they have not yet been adequately characterized in the human HF, we asked whether and where CIP/KIP proteins localise in the human hair matrix and pre-cortex in relation to cell cycle activity and HF-specific epithelial cell differentiation that is marked by keratin 85 (K85) protein expression. K85 expression coincided with loss or reduction in cell cycle activity markers, including in situ DNA synthesis (EdU incorporation), Ki-67, phospho-histone H3 and cyclins A and B1, affirming a post-mitotic state of pre-cortical HF keratinocytes. Expression of CIP/KIP proteins was found abundantly within the proliferative hair matrix, concomitant with a role in cell cycle checkpoint control. p21CIP1, p27KIP1 and cyclin E persisted within post-mitotic keratinocytes of the pre-cortex, whereas p57KIP2 protein decreased but became nuclear. These data imply a supportive role for CIP/KIP proteins in maintaining proliferative arrest, differentiation and anti-apoptotic pathways, promoting continuous hair bulb growth and hair shaft formation in anagen VI. Moreover, post-mitotic hair matrix regions contained cells with enlarged nuclei, and DNA in situ hybridisation showed cells that were >2N in the pre-cortex. This suggests that CIP/KIP proteins might counterbalance cyclin E to control further rounds of DNA replication in a cell population that has a propensity to become tetraploid. These data shed new light on the in situ-biography of human hair matrix keratinocytes on their path of active cell cycling, arrest and terminal differentiation, and showcase the human HF as an excellent, clinically relevant model system for cell cycle physiology research of human epithelial cells within their natural tissue habitat.


Correlation between eosinophils and protection against reinfection with Schistosoma mansoni and the effect of human immunodeficiency virus type 1 coinfection in humans.
Longitudinal investigations of an adult male population of Kenyan car washers who have heavy and quantifiable occupational exposure to Schistosoma mansoni cercariae revealed that some individuals develop resistance to reinfection while others remain highly susceptible. We sought to characterize immune correlates associated with host protection in this population. Previous studies have demonstrated an association of peripheral eosinophilia with resistance to reinfection with schistosomes. Thus, we investigated the relationship between the percentage of circulating eosinophils and the effect of human immunodeficiency virus type 1 (HIV-1) coinfection on the susceptibility of the car washers to reinfection with schistosomes. Elevated percentages of circulating eosinophils were associated with resistance to reinfection by S. mansoni in HIV-1-seronegative persons. In the HIV-1-seropositive cohort, low CD4+-T-cell counts were associated with a less intense eosinophilia. Moreover, eosinophils from the car washers expressed high levels of FcepsilonRI beta chain, a molecule important in immunoglobulin E (IgE)-mediated immunity. Levels of FcepsilonRI beta chain expression correlated with serum levels of total and antigen-specific IgE for HIV-1-negative car washers, but this was not the case for individuals coinfected with HIV-1. Overall, these data further implicate eosinophils as having a potential role in development of protective immunity against schistosomes and suggest that changes associated with HIV-1 coinfection increase susceptibility to reinfection.


Nanocomposite Hydrogels with Self-Assembling Peptide-Functionalized Carbon Nanostructures.
Carbon nanostructures (CNSs) are attractive components to attain nanocomposites, yet their hydrophobic nature and strong tendency to aggregate often limit their use in aqueous conditions and negatively impact their properties. In this work, carbon nanohorns (CNHs), multi-walled carbon nanotubes (CNTs), and graphene (G) are first oxidized, and then reacted to covalently anchor the self-assembling tripeptide L-Leu-D-Phe-D-Phe to improve their dispersibility in phosphate buffer, and favor the formation of hydrogels formed by the self-organizing L-Leu-D-Phe-D-Phe present in solution. The obtained nanocomposites are then characterized by transmission electron microscopy (TEM), oscillatory rheology, and conductivity measurements to gain useful insights as to the key factors that determine self-healing ability for the future design of this type of nanocomposites.


Characterization of an F1 deletion mutant of Yersinia pestis CO92, pathogenic role of F1 antigen in bubonic and pneumonic plague, and evaluation of sensitivity and specificity of F1 antigen capture-based dipsticks.
We evaluated two commercial F1 antigen capture-based immunochromatographic dipsticks, Yersinia Pestis (F1) Smart II and Plague BioThreat Alert test strips, in detecting plague bacilli by using whole-blood samples from mice experimentally infected with Yersinia pestis CO92. To assess the specificities of these dipsticks, an in-frame F1-deficient mutant of CO92 (Δcaf) was generated by homologous recombination and used as a negative control. Based on genetic, antigenic/immunologic, and electron microscopic analyses, the Δcaf mutant was devoid of a capsule. The growth rate of the Δcaf mutant generally was similar to that of the wild-type (WT) bacterium at both 26 and 37 °C, although the mutant's growth dropped slightly during the late phase at 37 °C. The Δcaf mutant was as virulent as WT CO92 in the pneumonic plague mouse model; however, it was attenuated in developing bubonic plague. Both dipsticks had similar sensitivities, requiring a minimum of 0.5 μg/ml of purified F1 antigen or 1 × 10(5) to 5 × 10(5) CFU/ml of WT CO92 for positive results, while the blood samples were negative for up to 1 × 10(8) CFU/ml of the Δcaf mutant. Our studies demonstrated the diagnostic potential of two plague dipsticks in detecting capsular-positive strains of Y. pestis in bubonic and pneumonic plague.


Cationic selectivity and competition at the sodium entry site in frog skin.
The cation selectivity of the Na entry mechanism located in the outer membrane of the bullfrog (Rana catesbeiana) skin epithelium was studied. This selectivity was determined by measuring the short-circuit current when all of the external sodium was replaced by another cation and, also, by noting the relative degree of inhibition that the alkali metal cations produced on Na influx. The ability of the Group Ia cations to permeate the apical membrane was determined from the tracer uptake experiments. The results demonstrate that (a) only Li and Na are actively transported through the epithelium; (b) the alkali cations K, Rb, and Cs do not enter the epithelium through the apical border and, therefore, Na and Li are the only alkali cations translocated through this membrane; (c) these impermeable cations are competitive inhibitors of Na entry; (d) the cations NH4 and Tl exhibit more complex behavior but, under well-defined conditions, also inhibit Na entry; and (e) the selectivity of the cation binding site is in the sequence Li congruent to Na > Tl > NH4 congruent to K > Rb > Cs, which corresponds to a high field strength site with tetrahedral symmetry.


The Effect of Acupuncture on Elbow Joint Sports Injuries Based on Magnetic Resonance Imaging.
Elbow joint injuries are extremely common in most athletes. Athletes' chronic elbow injuries can involve multiple complex anatomical structures related to orthopedics. Therefore, it is of great significance to explore the characteristics of magnetic resonance (MR) images of chronic sports injuries of the elbow joint and the influence of acupuncture treatment on MR images and clinical symptoms. A total of 60 elbow joints of 39 athletes from 15-25 years old were selected for coronal, sagittal, and axial MR scans to observe the image characteristics; 60 elbow joints were randomly divided into acupuncture group and control group and observed changes in MR images and clinical symptoms after acupuncture treatment. After acupuncture treatment, the clinical symptoms were significantly improved. MR images showed that the elbow joint effusion was reduced, and the bone marrow edema was reduced. The effective rate of acupuncture treatment is as high as 100%, while the effective rate of the control group is only 40%. Acupuncture treatment has significantly improved the range of motion of the elbow joint. Acupuncture treatment can significantly relieve athletes' elbow joint pain and locking symptoms, improve joint range of motion, and is beneficial to recovery of special training and high-level competitive competitions. It is a sensitive, effective, and noninvasive method.


Modern ECMO: why an ECMO programme in a tertiary care hospital.
Extracorporeal Membrane Oxygenation (ECMO) represents a useful tool to support the lungs and the heart when all conventional therapies failed and the patients are at risk of death. While the Extracorporeal Life Support Organization (ELSO) collects data from different institutions that joined the Registry and reports overall outcome, individual centres often collide with results below expectations, either in adults and in paediatric population. Some authors suggest that poor outcomes could be overcome with a programme dedicated to ECMO, with specialized professionals adequately trained on ECMO and with a consistent number of procedures. In 2012, The IRCCS PSD ECMO Programme was instituted with the specific aim of achieving better results than hitherto obtained. After only 1 year of activity, the results justified the programme, with a better survival rate for each group investigated, particularly in adults, but surprisingly in paediatrics too, where the results were better than what reported by ELSO. Although the number of patients treated with ECMO is still growing up, the effects of the ECMO programme continue to exert a positive action on outcome even now. The present article reports data on survival, blood loss, and blood consumption during ECMO in the last few years at our institution.


Alcohol drinking and cigarette smoking: a "partner" for gastric ulceration.
Alcohol consumption and cigarette smoking are two etiologic factors that have a close relationship with peptic ulcer diseases. Chronic active gastritis is reportedly associated with chronic alcohol ingestion. Nonetheless, the inflammatory changes are likely to be related to concurrent Helicobacter pylori infection that is common among alcoholics. Moreover, chronic alcoholism is also correlated with the presence of gastric metaplasia. Both clinically and experimentally, alcohol had been shown to affect the mucosal barrier and histology. These ulcerogenic effects play a crucial role in altering gastric mucosal defense mechanisms. Cigarette smoking is coupled with the initiation and prolongation of gastric ulcers. Epidemiologic data show that cigarette smoking increases both the incidence and relapse rate of peptic ulcer diseases and also delays ulcer healing in humans. Retrospective studies also indicate that cigarette smoking is a key factor in inducing ulcer diseases rather than a linked behavior. The general detrimental effects of cigarette smoking in the gastric mucosa include reduction of circulating epidermal growth factor, increase in tissue free radical production and the presence of free radicals in smoke, together with reduction of mucosal constitutive nitric oxide synthase activity. Furthermore, the alteration of normal gastric mucosal blood flow and angiogenesis and the suppression of cell proliferation contribute largely to the delay in ulcer healing in cigarette smokers. Concurrent consumption of alcohol and cigarette smoking significantly increases the risk of gastric ulcers. In animal experiments, cigarette smoking potentiated ethanol-induced gastric mucosal damage. The reduction of mucus secretion, increase in leukotriene B4 level, increased activities of inducible nitric oxide synthase, xanthine oxidase and myeloperoxidase, and the expression of adhesion molecules in the gastric mucosa accompanied such potentiating effects. Substances other than nicotine in cigarette smoke may also contribute to the above effects.


Silencing of Euchromatic Transposable Elements as a Consequence of Nuclear Lamina Dysfunction.
Transposable elements (TEs) are mobile genomic sequences that are normally repressed to avoid proliferation and genome instability. Gene silencing mechanisms repress TEs by RNA degradation or heterochromatin formation. Heterochromatin maintenance is therefore important to keep TEs silent. Loss of heterochromatic domains has been linked to lamin mutations, which have also been associated with derepression of TEs. In fact, lamins are structural components of the nuclear lamina (NL), which is considered a pivotal structure in the maintenance of heterochromatin domains at the nuclear periphery in a silent state. Here, we show that a lethal phenotype associated with Lamin loss-of-function mutations is influenced by Drosophilagypsy retrotransposons located in euchromatic regions, suggesting that NL dysfunction has also effects on active TEs located in euchromatic loci. In fact, expression analysis of different long terminal repeat (LTR) retrotransposons and of one non-LTR retrotransposon located near active genes shows that Lamin inactivation determines the silencing of euchromatic TEs. Furthermore, we show that the silencing effect on euchromatic TEs spreads to the neighboring genomic regions, with a repressive effect on nearby genes. We propose that NL dysfunction may have opposed regulatory effects on TEs that depend on their localization in active or repressed regions of the genome.


Fatty Liver Index as a proxy for non-alcoholic fatty liver disease and the risk of stroke and coronary heart disease: The Suita Study.
Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in developed countries, but its role in predicting cardiovascular disease (CVD) needs further investigation. Herein, we studied the association between NAFLD and the risk of CVD, stroke, and coronary heart disease (CHD) among Japanese people. This prospective cohort study analyzed data from 2,517 men and 3,958 women, aged 30-84 years, who were registered in the Suita Study. NAFLD was defined as Fatty Liver Index (FLI) ≥ 60. Cox proportional hazard models were applied to calculate the hazard ratios (HRs) and 95% confidence intervals (95% CIs) of incident CVD, stroke, and CHD events by baseline FLI. The results were adjusted for age, smoking, alcohol consumption, hypertension, diabetes, lipid profile, chronic kidney disease, and cardiac murmur or valvular diseases. Within 16.6 years of median follow-up, 590 participants developed CVD (346 stroke events and 244 CHD events). Women with NAFLD (FLI ≥ 60) showed a higher risk of CVD and stroke: HRs (95% CIs) = 1.69 (1.16, 2.46) and 2.06 (1.31, 3.24), respectively. Besides, women in the fourth and fifth (highest) FLI quintiles showed a higher risk of CVD and stroke than those in the third (middle) quintile: HRs (95% CIs) = 1.60 (1.08, 2.36) and 1.67 (1.13, 2.45) for CVD and 1.73 (1.07, 2.79) and 1.90 (1.18, 3.05) for stroke, respectively. No corresponding associations were detected in men. NAFLD was not associated with CHD risk in either sex. NAFLD, diagnosed by FLI, was associated with a higher risk of CVD and stroke in Japanese women. From a preventive perspective, women with NAFLD should be targeted for CVD screenings and interventions.


Purification, biochemical properties and active sites of N-acetyl-beta-D-hexosaminidases from human seminal plasma.
Two isoenzymes of N-acetyl-beta-D-glucosaminidase (EC 188.8.131.52) (Hex A and Hex B) from human seminal plasma were purified to homogeneity with specific activities of 26 and 60 units/mg of protein respectively. N-Acetyl-beta-D-glucosaminidase activity was inseparable from N-acetyl-beta-D-galactosaminidase activity in both Hex A and Hex B by various conventional chromatographic procedures. Although Km values of N-acetyl-beta-glucosaminidase activity of Hex A and Hex B were similar (1.33 mM), those of N-acetyl-beta-galactosaminidase activity were 0.14 mM for Hex A and 0.40 mM for Hex B. However, pH optima and temperature optima were identical for N-acetyl-beta-glucosaminidase and N-acetyl-beta-galactosaminidase activities of both isoenzymes; Hex A was far more heat-sensitive than Hex B. Thiol-reactive compounds such as silver salts, mercuric salts, p-chloromercuribenzoate and thimerosal strongly inhibited the N-acetyl-beta-glucosaminidase activities of both isoenzymes. GSH protected the enzyme activities from inactivation caused by these reagents, confirming the presence of thiol groups at the active centres. Inhibitions of N-acetyl-beta-glucosaminidase activities of both isoenzymes by metal salts and organic anions were comparable; acetate and arsenite were effective inhibitors for both isoenzymes. In contrast, inhibitions of N-acetyl-beta-glucosaminidase activities of the two isoenzymes by iodoacetic acid, iodoacetamide and ethylmaleimide were not comparable; Hex B was more susceptible to inhibition by these agents at 20 mM concentration. The N-acetyl-beta-glucosaminidase activities of both isoenzymes are strongly inhibited, in decreasing order, by N-acetyl-galactosamine, mannosamine, disaccharic acid lactone, N-acetylglucosamine and gluconolactone. The Ki values of the N-acetyl-beta-glucosaminidase and N-acetyl-beta-galactosaminidase activities for N-acetylhexosamines and results from mixed-substrate kinetics indicated that the activities for the two substrates are located at different sites in Hex A and at the same site in Hex B. The Mr values of Hex A and Hex B were determined to be 195,000 and 210,000 respectively by gel filtration through Sephadex G-200. SDS/polyacrylamide-gel electrophoresis revealed that Hex A and Hex B are each composed of four subunits corresponding to Mr about 50,000 each. No further polypeptide chain was obtained after reduction and alkylation of Hex A and Hex B with 10 mM-dithiothreitol and 10 mM-iodoacetamide.


Symptom clusters for revising scale membership in the analysis of prostate cancer patient reported outcome measures: a secondary data analysis of the Medical Research Council RT01 trial (ISCRTN47772397).
To investigate the role of symptom clusters in the analysis and utilisation of patient reported outcome measures (PROMs) for data modelling and clinical practice. To compare symptom clusters with scales, and to explore their value in PROMs interpretation and symptom management. A dataset called RT01 (ISCRTN47772397) of 843 prostate cancer patients was used. PROMs were reported with the University of California, Los Angeles Prostate Cancer Index (UCLA-PCI). Symptom clusters were explored with hierarchical cluster analysis (HCA) and average linkage method (correlation > 0.6). The reliability of the Urinary Function Scale was evaluated with Cronbach's Alpha. The strength of the relationship between the items was investigated with Spearman's correlation. Predictive accuracy of the clusters was compared to the scales by receiver operating characteristic (ROC) analysis. Presence of urinary symptoms at 3 years measured with the late effects on normal tissue: subjective, objective, management tool (LENT/SOM) was an endpoint. Two symptom clusters were identified (urinary cluster and sexual cluster). The grouping of symptom clusters was different than UCLA-PCI Scales. Two items of the urinary function scales ("number of pads" and "urinary leak interfering with sex") were excluded from the urinary cluster. The correlation with the other items in the scale ranged from 0.20 to 0.21 and 0.31 to 0.39, respectively. Cronbach's Alpha showed low correlation of those items with the Urinary Function Scale (0.14-0.36 and 0.33-0.44, respectively). All urinary function scale items were subject to a ceiling effect. Clusters had better predictive accuracy, AUC = 0.70 -0.65, while scales AUC = 0.67-0.61. This study adds to the knowledge on how cluster analysis can be applied for the interpretation and utilisation of PROMs. We conclude that multiple-item scales should be evaluated and that symptom clusters provide a study-specific approach for modelling and interpretation of PROMs.


Incidence rates of acute promyelocytic leukemia among Hispanics, blacks, Asians, and non-Hispanic whites in the United States.
Despite significant improvements in the prognosis of acute promyelocytic leukemia brought about by therapeutic advances, understanding of the epidemiology of acute promyelocytic leukemia remains limited. Earlier reports have suggested that Hispanics may have an increased incidence of acute promyelocytic leukemia, but no systematic analysis of national data has yet been reported. We performed a retrospective cohort study, using data from the Surveillance, Epidemiology, and End Results (SEER) Program of the National Cancer Institute from 1992-2001 in order to compare leukemia incidence rates as a function of race and ethnicity. We identified 709 cases of acute promyelocytic leukemia and analyzed incidence rates by race and sex. Hispanics were not found to have greater lifetime incidence rates than whites, with an incidence relative rate (IRR) of 0.86 that of whites (P=0.17). The age distribution among Hispanics was significantly different from non-Hispanic whites, with greater incidence rates for children ages 1-19 years (IRR=1.9, P=0.02) and adult ages 20-44 years (IRR=1.6, P=0.004). Blacks had lower lifetime incidence rates than non-Hispanic whites (IRR=0.75, P=0.04), Hispanics (IRR=0.64, P=0.007), and Asians (IRR=0.67, P=0.03). Asians did not differ from non-Hispanic whites in lifetime or age-specific incidence rates. These results indicate that while US Hispanics do not have greater lifetime incidence rates of acute promyelocytic leukemia, blacks have lower incidence rates of acute promyelocytic leukemia than Hispanics, non-Hispanic whites, and Asians.


Inhibition of Na+/K+-ATPase by 5,6,7,8-tetrafluoro-3-hydroxy-2-phenylquinolin-4(1H)-one.
Na+/K+-ATPase (NKA) is an enzyme of crucial importance for all animal cells. We examined the inhibitory effects of halogenated phenylquinolinones on NKA. The 5,6,7,8-tetrafluoro-3-hydroxy-2-phenylquinolin-4(1H)-one (TFHPQ) was identified as an efficient NKA inhibitor with IC50 near 10 μM. The inhibition by TFHPQ is particularly efficient at higher concentrations of K+, where NKA adopts the E2 conformation. The experimental observations are in a good agreement with the outcomes from molecular docking. We identified an energetically favourable TFHPQ binding site for the K+-bound NKA, which is located in the proximity of the cytoplasmic C-terminus.


The association of multimorbidity within cardio-metabolic disease domains with dietary patterns: A cross-sectional study in 129 369 men and women from the Lifelines cohort.
Multimorbidity is considered a major challenge for current health care. Lifestyle interventions, as a broad and generic approach, may have the potential to improve the management of care among patients with multimorbidity. The objective of this study was to evaluate the association of multimorbidity defined within the cardiometabolic disease domains with dietary patterns, representing habitual dietary intake. We studied 129 369 participants from the Lifelines Cohort study (42% male, 45±13 years (range 18-93)) in which diet was assessed using a 110-item food frequency questionnaire. A composite morbidity score was applied in multivariable ordered logistic regression to test the association with dietary patterns derived by principal components analysis, based on sex-specific dietary pattern scores. Four dietary patterns were retained, accounting for 26.6% of the variation in overall diet. After control for potential confounders, men and women in the highest quintile of "meat, alcohol and potato pattern" and "snack pattern" had a higher likelihood of having higher morbidity scores than those in the lowest quintile (e.g. men: OR = 1.83(95% CI:1.71-1.97), OR = 1.18(95% CI 1.11-1.27 respectively). The opposite was observed with respect to the "bread and sweets pattern" and "vegetable, fish and fruit pattern" (e.g. women: OR = 0.88(95% CI: 0.81-0.96), OR = 0.86(95% CI 0.81-0.92 respectively). The association partially attenuated after adjusting for BMI, but the associations remained significant among men. Robust associations between dietary patterns and multimorbidity within the cardiometabolic domain, in particular a "meat, alcohol and potato pattern", suggest an important opportunity of dietary interventions in multimobidity prevention. Generic prevention strategies based on population derived dietary patterns may have the potential to enhance lifestyle management among people with multimorbidity.


Intravesical delivery of rapamycin suppresses tumorigenesis in a mouse model of progressive bladder cancer.
Early-stage bladder cancer occurs as two distinct forms: namely, low-grade superficial disease and high-grade carcinoma in situ (CIS), which is the major precursor of muscle-invasive bladder cancer. Although the low-grade form is readily treatable, few, if any, effective treatments are currently available for preventing progression of nonmuscle-invasive CIS to invasive bladder cancer. Based on our previous findings that the mammalian target of Rapamycin (mTOR) signaling pathway is activated in muscle-invasive bladder cancer, but not superficial disease, we reasoned that suppression of this pathway might block cancer progression. To test this idea, we performed in vivo preclinical studies using a genetically engineered mouse model that we now show recapitulates progression from nonmuscle-invasive CIS to muscle-invasive bladder tumors. We find that delivery of Rapamycin, an mTOR inhibitor, subsequent to the occurrence of CIS effectively prevents progression to invasive bladder cancer. Furthermore, we show that intravesical delivery of Rapamycin directly into the bladder lumen is highly effective for suppressing bladder tumorigenesis. Thus, our findings show the potential therapeutic benefit of inhibiting mTOR signaling for treatment of patients at high risk of developing invasive bladder cancer. More broadly, our findings support a more widespread use of intravesical delivery of therapeutic agents for treatment of high-risk bladder cancer patients, and provide a mouse model for effective preclinical testing of potential novel agents.


Modified nanofat grafting: Stromal vascular fraction simple and efficient mechanical isolation technique and perspectives in clinical recellularization applications.
Background: Nanofat grafting (NG) is a simple and cost-effective method of lipoaspirates with inter-syringe passages, to produce stromal vascular fraction (SVF) and isolate adipose-derived stem cells (ASCs). This represents a tremendous interest in the future clinical needs of tissue engineering. In this study, we optimized the NG technique to increase the yield of ASC extractions. Methods: We analyzed three groups of SVF obtained by 20, 30, and 40 inter-syringe passages. The control group was an SVF obtained by enzymatic digestion with Celase. We studied their cell composition by flow cytometry, observed their architecture by confocal microscopy, and observed immunomodulatory properties of the ASCs from each of the SVFs by measuring inflammatory markers of macrophages obtained by an ASC monocyte co-culture. Results: We have established the first cell mapping of the stromal vascular fraction of adipose tissue. The results showed that SVF obtained by 20 inter-syringe passages contains more statistically significant total cells, more cells expressing the ASC phenotype, more endothelial cells, and produces more CFU-F than the SVF obtained by 30 and 40 passages and by enzymatic digestion. Confocal microscopy showed the presence of residual adipocytes in SVF obtained by inter-syringe passages but not by enzymatic digestion. The functional study indicates an orientation toward a more anti-inflammatory profile and homogenization of their immunomodulatory properties. Conclusion: This study places mechanically dissociated SVF in the center of approaches to easily extract ASCs and a wide variety and number of other progenitor cells, immediately available in a clinical setting to provide both the amount and quality of cells required for decellularized tissues.


Immunological heterogeneity of human monocyte subsets prepared by counterflow centrifugation elutriation.
Human peripheral blood mononuclear cells were separated into three fractions by means of counterflow centrifugation elutriation (CCE). The first fraction, eluted at a flow rate of 24 ml/min, was composed of lymphocytes with less than 2% contaminating esterase-positive cells. The cells in this fraction were incapable of responding to either soluble antigen (tetanus toxoid) or particulate antigen (cytomegalovirus-infected fibroblasts) unless recombined with accessory cells. The second fraction, eluted at a flow rate of 28 ml/min, was composed predominantly (72%) of small Ia, leu M3, and esterase-positive monocytes, which stained weakly with leu 10 antibody. Cells in this fraction efficiently presented soluble and particulate antigens to monocyte-depleted lymphocytes. Of the remaining cells, 87% were large esterase-positive monocytes that labelled strongly with Ia, leu M3, and leu 10. These cells were less efficient in antigen presentation than the small monocytes. However, lymphocytes activated with antigen-pulsed large monocytes exhibited more suppressor cell activity than those activated with antigen-pulsed small monocytes.


Setup and Troubleshooting of Volta Phase Plate Cryo-EM Data Collection.
Cryo electron microscopy (cryo-EM) has become a method of choice in structural biology to analyze isolated complexes and cellular structures. This implies adequate imaging of the specimen and advanced image-processing methods to obtain high-resolution 3D reconstructions. The use of a Volta phase plate in cryo-EM drastically increases the image contrast while being able to record images at high acceleration voltage and close to focus, i.e., at conditions where high-resolution information is best preserved. During image processing, higher contrast images can be aligned and classified better than lower quality ones resulting in increased data quality and the need for less data. Here, we give step-by-step guidelines on how to set up high-quality VPP cryo-EM single particle data collections, as exemplified by human ribosome data acquired during a one-day data collection session. Further, we describe specific technical details in image processing that differ from conventional single particle cryo-EM data analysis.


Sakyomicins A, B, C and D: new quinone-type antibiotics produced by a strain of Nocardia. Taxonomy, production, isolation and biological properties.
Actinomycete strain M-53, a new soil isolate, was found to produce four quinone-type antibiotics. Antibiotic sakyomicin components, A, B, C and D were isolated from the fermentation broth of strain M-53 by XAD-2 column chromatography, silica gel column chromatography and Sephadex LH-20 column chromatography. The components are active against Gram-positive bacteria. Strain M-53 was identified as a strain of genus Nocardia.


Increase in HIV diagnoses among men who have sex with men in New Zealand from a stable low period.
To describe trends in HIV diagnoses among men who have sex with men (MSM) in New Zealand 1996-2008, and to identify characteristics associated with HIV diagnoses in the resurgent phase. Data collected through routine surveillance of HIV infection, where the mode of transmission included homosexual contact, were analysed over the period 1996-2008. Annual HIV diagnoses were low during 1996-2000, rose sharply between 2001 and 2005, and remained at an elevated plateau between 2006 and 2008. Over a quarter were attributed to HIV infection acquired overseas (28.6%). Trends in diagnoses of locally acquired HIV infection closely mirrored the trend of three diagnosis phases. Increases in locally acquired HIV occurred among virtually all characteristics of MSM. However, compared with MSM diagnosed in the low phase 1996-2000, individuals diagnosed in the resurgent phase 2001-05 were more likely to be aged 30-39, to have tested HIV-negative within the previous 2 years, to live in the Northern region encompassing Auckland, and to be of non-European ethnicity. The per capita HIV diagnosis rate among MSM was lowest in 1997, at 22.0 per million males aged 15-64, and highest in 2005 at 66.7 per million. The increase in HIV diagnoses among MSM in New Zealand was primarily due to an increase in locally acquired HIV infection, which disproportionately affected some groups of MSM. Factors driving this change in local epidemic conditions need to be identified. The rate of new HIV diagnoses among MSM remains low by international standards.


Efficacy and safety of anticoagulation in non-malignant portal vein thrombosis in patients with liver cirrhosis: a systematic review and meta-analysis.
The role of anticoagulation in treating non-malignant portal vein (PV) thrombosis (PVT) in patients with liver cirrhosis remains unclear. In our meta-analysis, we aimed to evaluate the efficacy and safety of anticoagulation for the treatment of non-malignant PVT in these patients. We conducted a meta-analysis to estimate the effects of anticoagulation on non-malignant PVT in patients with liver cirrhosis. We assessed the rates of PV recanalization, variceal bleeding, and any bleeding. We included 9 observational studies which involved 474 patients. The rate of PV recanalization was significantly higher in patients who received anticoagulation compared to those who did not: 65.2% vs. 25.2%; relative risk (RR) 2.31, 95% confidence interval (CI) 1.80-2.96; P<0.00001. Variceal bleeding was significantly lower in patients who received anticoagulation: 0.1% vs. 18.5%; RR 0.15, 95%CI 0.04-0.55; P=0.004. Any bleeding was similar between patients who received anticoagulation and those who did not: 10.3% vs. 22.7%; RR 0.43, 95%CI 0.09-1.99; P=0.28. Anticoagulation use increased the rate of PV recanalization in cirrhotic patients with non-malignant PVT. Anticoagulation decreased the rate of variceal bleeding and did not increase the rate of any bleeding.


Differential expression of cyclins A, B1, D3 and E in G1 phase of the cell cycle between the synchronized and asynchronously growing MOLT-4 cells.
The use of 'double-thymidine block' was the first widely accepted method for inducing cell synchrony and remains one of the most effective and frequently used techniques for analyzing the cell cycle today. While thymidine is in itself an inhibitor of DNA replication, thymidine blocks are typically used to generate cell synchrony at the G1/S boundary. We have previously presented the first evidence that shows the growth imbalance and altered expression levels of cyclins A, B1, D3 and E in MOLT-4 cells synchronized in the cell cycle by thymidine. The major objective of the present study was to compare the levels of cyclins A, B1, D3 and E in G1 phase of the cell cycle between synchronized and unperturbed asynchronously growing human lymphocyte leukemia MOLT-4 cells. Here, we demonstrate that the sorted, asynchronously growing MOLT-4 cells had considerably lower levels of cyclins A, B1, D3 and E than their counterparts of the cells arrested in G1/S phase, as assessed by flow cytometry. In addition, we confirmed these results by using post-sorting Western blotting, a new method we recently developed for examining protein expression in specific phases of sorted, synchronized or asynchronously growing cells. Our findings revealed that the levels of cyclins D3 and E in the asynchronously growing MOLT-4 cells were significantly lower than those in synchronized cultures. Interestingly, protein expression levels of cyclins A and B1 in the asynchronously growing MOLT-4 cells were barely measurable, suggesting that these proteins were either not expressed or under detectable levels. These studies indicate that our synchronization protocol may have disturbed cell proliferation and metabolism as evidenced by significant differences in the expression of cyclins between asynchronously growing and synchronized cells, and further suggest that the levels of cyclins A, B1, D3 and E in synchronized cultures cannot represent those in unperturbed, asynchronously growing cells. Thus, it appears that thymidine-treated, synchronized cells may not be suitable experimental models for the study of normal cell cycle.


Circulating and excreted hormones during the ovarian cycle in the cotton-top tamarin, Saguinus oedipus.
Five parous, cycling cotton-top tamarin females were used to compare circulating and urinary profiles of the major reproductive hormones during the ovarian cycle. Blood samples were collected on unanesthetized, hand restrained females three times per week for 25 days and first morning void urine was collected daily for 30-40 days including the serum sampling period. Serum and urine samples were analyzed for estrone, estradiol, and bioactive LH. Additionally, serum was analyzed for progesterone and urine for estrone conjugates (E1G) and immunoreactive LH. Both serum and urinary hormonal profiles revealed ovarian cyclicity in four females; the fifth one stopped cycling during the study. All hormones were referenced to the urinary immunoreactive LH peak. Urinary estrone conjugates followed similar profiles to serum progesterone. Progesterone levels increased two- to threefold on the day of the urinary immunoreactive LH peak. Peak levels of circulating bioactive LH occurred on the same day or the day preceding the urinary bioactive and immunoreactive LH peak for all four cycling females. Serum estradiol levels rose during the follicular phase and peaked prior to or on the day of the urinary LH peak while urinary estradiol levels did not rise until the day of the urinary LH peak. Serum and urinary estrone levels did not rise until the day of the urinary LH peak and remained high throughout the luteal phase. The measurement of urinary LH corresponded well with serum LH, establishing urinary LH as an important hormone for monitoring the ovulation in the tamarin. The highest circulating estradiol concentrations were found in the follicular phase of the cycle, but estradiol did not appear in urine during the follicular phase of the cycle. Elevated estrone levels occur during the luteal phase following the progesterone pattern, and therefore elevated urinary concentrations of estrone and estrone conjugates may be indicative of luteal function. © 1993 Wiley-Liss, Inc.


Atomistic studies of surface adhesions using molecular-dynamics simulations.
The existing theoretical descriptions of continuum surface adhesions, such as the JKR (Johnson-Kendall-Roberts) model, have been very useful for the interpretation of particle contacts of sizes down to micrometre length-scales. However, the continuum model is expected to fail at atomic length-scales, where discrete atomistic interactions become significant. The crossover length-scales at which the macroscopic phenomena begin to manifest themselves, and how they occur, are equally baffling and remain poorly understood. This paper explores the issue of atomistic surface contacts between two ionic bodies made of similar materials, such as MgO and NaCl, using molecular dynamics. A range of surface phenomenological behaviour has been identified, from simple fracture to plastic dislocation and neck formation, which are not predicted in continuum models. In addition, the influences of body structure with respect to surface contacts will also be discussed briefly.


Perceptual grouping and visual enumeration.
We used lateralized Event-Related Potential (ERP) measures - the N2pc and CDA/SPCN components - to assess the role of grouping by target similarity during enumeration. Participants saw a variable number (0, 1, 2 or 3) of same- or differently-colored targets presented among homogeneous distracters, and performed an enumeration task. Results showed that the N2pc, but not the CDA, was larger for multiple targets of identical color relative to targets of different colors. The findings are interpreted in terms of the effects of grouping on early versus late stages of multiple object processing. Within this framework, they reveal that grouping has an effect on early individuation mechanisms, while later processing mechanisms are less prone to such an influence.


Chest CT Computerized Aided Quantification of PNEUMONIA Lesions in COVID-19 Infection: A Comparison among Three Commercial Software.
To compare different commercial software in the quantification of Pneumonia Lesions in COVID-19 infection and to stratify the patients based on the disease severity using on chest computed tomography (CT) images. We retrospectively examined 162 patients with confirmed COVID-19 infection by reverse transcriptase-polymerase chain reaction (RT-PCR) test. All cases were evaluated separately by radiologists (visually) and by using three computer software programs: (1) Thoracic VCAR software, GE Healthcare, United States; (2) Myrian, Intrasense, France; (3) InferRead, InferVision Europe, Wiesbaden, Germany. The degree of lesions was visually scored by the radiologist using a score on 5 levels (none, mild, moderate, severe, and critic). The parameters obtained using the computer tools included healthy residual lung parenchyma, ground-glass opacity area, and consolidation volume. Intraclass coefficient (ICC), Spearman correlation analysis, and non-parametric tests were performed. Thoracic VCAR software was not able to perform volumes segmentation in 26/162 (16.0%) cases, Myrian software in 12/162 (7.4%) patients while InferRead software in 61/162 (37.7%) patients. A great variability (ICC ranged for 0.17 to 0.51) was detected among the quantitative measurements of the residual healthy lung parenchyma volume, GGO, and consolidations volumes calculated by different computer tools. The overall radiological severity score was moderately correlated with the residual healthy lung parenchyma volume obtained by ThoracicVCAR or Myrian software, with the GGO area obtained by the ThoracicVCAR tool and with consolidation volume obtained by Myrian software. Quantified volumes by InferRead software had a low correlation with the overall radiological severity score. Computer-aided pneumonia quantification could be an easy and feasible way to stratify COVID-19 cases according to severity; however, a great variability among quantitative measurements provided by computer tools should be considered.