Non-antigen-contacting region of an asymmetric bispecific antibody to factors IXa/X significantly affects factor VIII-mimetic activity.
While antibody engineering improves the properties of therapeutic antibodies, optimization of regions that do not contact antigens has been mainly focused on modifying the effector functions and pharmacokinetics of antibodies. We recently reported an asymmetric anti-FIXa/FX bispecific IgG4 antibody, ACE910, which mimics the cofactor function of FVIII by placing the two factors into spatial proximity for the treatment of hemophilia A. During the optimization process, we found that the activity was significantly affected by IgG subclass and by modifications to the inter-chain disulfide bonds, upper hinge region, elbow hinge region, and Fc glycan, even though these regions were unlikely to come into direct contact with the antigens. Of these non-antigen-contacting regions, the tertiary structure determined by the inter-chain disulfide bonds was found to strongly affect the FVIII-mimetic activity. Interestingly, IgG4-like disulfide bonds between Cys131 in the heavy chain and Cys114 in the light chain, and disulfide bonds between the two heavy chains at the hinge region were indispensable for the high FVIII-mimetic activity. Moreover, proline mutations in the upper hinge region and removal of the Fc glycan enhanced the FVIII-mimetic activity, suggesting that flexibility of the upper hinge region and the Fc portion structure are important for the FVIII-mimetic activity. This study suggests that these non-antigen-contacting regions can be engineered to improve the biological activity of IgG antibodies with functions similar to ACE910, such as placing two antigens into spatial proximity, retargeting effector cells to target cells, or co-ligating two identical or different antigens on the same cell.


Synthesis and In Vitro and In Vivo Evaluation of [3H]LRRK2-IN-1 as a Novel Radioligand for LRRK2.
LRRK2 (leucine-rich repeat kinase 2) has recently been proven to be a promising drug target for Parkinson's disease (PD) due to an apparent enhanced activity caused by mutations associated with familial PD. To date, there have been no reports in which a LRRK2 inhibitor has been radiolabeled and used for in in vitro or in vivo studies of LRRK2. In the present study, we radiolabeled the LRRK2 ligand, LRRK-IN-1, for the purposes of performing in vitro (IC50, K d , B max, autoradiography) and in vivo (biodistribution, and blocking experiments) evaluations in rodents and human striatum tissues. [3H]LRRK2-IN-1 was prepared with high radiochemical purity (>99 %) and a specific activity of 41 Ci/mmol via tritium/hydrogen (T/H) exchange using Crabtree's catalyst. For IC50, K d , and B max determination, LRRK2-IN-1 was used as a competing drug for nonspecific binding assessment. The specific binding of the tracer was further evaluated via an in vivo blocking study in mice with a potent LRRK2 inhibitor, Pf-06447475. In vitro binding studies demonstrated a saturable binding site for [3H]LRRK2-IN-1 in rat kidney, rat brain striatum and human brain striatum with K d of 26 ± 3 and 43 ± 8, 48 ± 2 nM, respectively. In rat, the density of LRRK2 binding sites (B max) was higher in kidney (6.4 ± 0.04 pmol/mg) than in brain (2.5 ± 0.03 pmol/mg), however, in human brain striatum, the B max was 0.73 ± 0.01 pmol/mg protein. Autoradiography imaging in striatum of rat and human brain tissues gave results consistent with binding studies. In in vivo biodistribution and blocking studies in mice, co-administration with Pf-06447475 (10 mg/kg) reduced the uptake of [3H]LRRK2-IN-1 (%ID/g) by 50-60% in the kidney or brain. The high LRRK2 brain density observed in our study suggests the feasibility for positron emission tomography imaging of LRRK2 (a potential target) with radioligands of higher affinity and specificity.


In a search for a shape maximizing packing fraction for two-dimensional random sequential adsorption.
Random sequential adsorption of various two dimensional objects is studied in order to find a shape which maximizes the saturated packing fraction. This investigation was begun in our previous paper [Cieśla et al., Phys. Chem. Chem. Phys. 17, 24376 (2015)], where the densest packing was studied for smoothed dimers. Here this shape is compared with the smoothed n-mers, spherocylinders, and ellipses. It is found that the highest packing fraction out of the studied shapes is 0.584 05 ± 0.0001 and is obtained for ellipses having long-to-short axis ratio of 1.85 ± 0.07.


Characterization of microenvironments of 2-(4'-hydroxyphenylazo)benzoic acid bound to bovine serum albumin by studying the solvent effects.
The binding and thermodynamic parameters of the two binding forms of 2-(4'-hydroxyphenylazo)benzoic acid to bovine serum albumin were estimated. The number of the binding sites of the azo form was 3.2 and that of the hydrazone form was 0.7. The binding of the hydrazone form was affected only by enthalpy changes, in contrast to that of the azo form which was affected by both enthalpy and entropy changes. To estimate the complex microenvironments of 2-(4'-hydroxyphenylazo)benzoic acid molecules on bovine serum albumin, the solvent effects were studied. The 2-carboxyl group of 2-(4'-hydroxyphenylazo)benzoic acid participates in the azo-hydrazone tautomerism . The 2-carboxylate ion forms an ion pair with triethylamine in ethylene dichloride, chloroform, and benzene, resulting in the appearance of the hydrazone form. The hydrazone formation in the system of 2-(4'-hydroxyphenylazo)benzoic acid-triethylamine (1:1) in chloroform was affected only by enthalpy changes, in the same manner as in the system of 2-(4'-hydroxyphenylazo)benzoic acid-bovine serum albumin. We speculate the presence of the two kinds of ion pairs on the basis of the changes of the azo-hydrazone tautomerism in chloroform, and that the azo form takes the contact ion pair and the hydrazone form takes the solvent-separated ion pair. A new possible model for the interaction of the azo and hydrazone forms and bovine serum albumin is proposed.


Type IV collagen aggregates promote keratinocyte proliferation and formation of epidermal layer in human skin equivalents.
Type IV collagen isolated from lens capsule without enzymatic treatment is known to form a gel under physiological condition and influences cellular activities. In case of human keratinocytes, the suppression of proliferation on reconstituted type IV collagen gels was reported in monolayer culture. In this study, we examined effects of type IV collagen isolated from porcine lens capsule on epidermal formation in human skin equivalents (HSEs). Type IV collagen aggregates were prepared under the culture condition and the aggregates suppressed keratinocyte proliferation in monolayer culture as well as the culture on the gels. In HSEs, type IV collagen aggregates were reconstituted on the surface of contracted collagen gels containing human dermal fibroblasts and the keratinocytes were then cultured on the aggregates for 14 days. Interestingly, in HSEs with type IV collagen aggregates, the BrdU-positive keratinocytes were increased and the thickness of the epidermal layer was around twice than that of control culture. Epidermal differentiation markers were expressed in the upper layer of the epidermis and the defined deposition of human basement membrane components were increased at the dermal-epidermal junction. These results indicate that the type IV collagen aggregates stimulate the proliferation of basal keratinocytes and improve the stratification of epidermal layers in HSEs.


HIV and STI prevalence and risk factors among male sex workers and other men who have sex with men in Nairobi, Kenya.
: Previous surveys of men who have sex with men (MSM) in Africa have not adequately profiled HIV status and risk factors by sex work status. MSM in Nairobi, Kenya, were recruited using respondent-driven sampling, completed a behavioral interview, and were tested for HIV and sexually transmitted infections. Overlapping recruitment among 273 male sex workers and 290 other MSM was common. Sex workers were more likely to report receptive anal sex with multiple partners (65.7% versus 18.0%, P < 0.001) and unprotected receptive anal intercourse (40.0% versus 22.8%, P = 0.005). Male sex workers were also more likely to be HIV infected (26.3% versus 12.2%, P = 0.007).


Vanilloid receptors--do they have a role in whole body metabolism? Evidence from TRPV1.
With increasing lifespan, therapeutic interventions for the treatment of disorders such as type 2 diabetes mellitus are in great demand. Despite billions of dollars invested to reduce the symptoms and complications due to diabetes mellitus, current treatments (e.g., insulin replacements, sensitization) remain inadequate, justifying the search for novel therapeutic approaches or alternative solutions, including dietary supplementation, for the treatment of diabetes mellitus in every age group. The involvement of the vanilloid system in the regulation of metabolism has been identified, and the emerging role of its receptors, the transient receptor potential vanilloid type 1 (TRPV1), in diabetes was recently demonstrated. Indeed, beneficial effects of dietary capsaicin, an agonist of TRPV1 receptors, were identified for improving glucose, insulin and glucagon-like peptide-1 levels. Recent findings regarding TRPV1 receptors in association with whole body metabolism including glucose homeostasis will be reviewed in this article.


Human fibroblast interferon gene lacks introns.
A recombinant lambda bacteriophage isolated from a human genome library contains the gene for fibroblast interferon (IFN-beta1). The DNA sequence of this gene is identical to the sequence of its mRNA and is devoid of introns.


1-Methylcyclopropene affects the antioxidant system of apricots (Prunus armeniaca L. cv. Búlida) during storage at low temperature.
Apricots (Prunus armeniaca cv. Búlida) were treated with 1 μL L⁻¹ [corrected] 1-methylcyclopropene (1-MCP) immediately after harvest and stored in air at 2 degrees C for 21 days. Antioxidant levels (ascorbic acid and carotenoids), enzymatic antioxidant activities (superoxide dismutase (SOD) and unspecific peroxidase (POX)) and total antioxidant capacity (trolox equivalent antioxidant capacity (TEAC)) were determined. The level of oxidative stress was also established by measuring ion leakage during storage. The changes in the antioxidant potential of apricots were related to the capacity of 1-MCP to increase their commercial life. 1-MCP-treated fruits exhibited higher SOD activity, whereas POX activity was significantly higher only after 21 days at 2 degrees C. Treated fruits also exhibited better retention of ascorbate and carotenoids and higher TEAC during storage. In accordance with these observations, lower ion leakage values were detected in 1-MCP-treated apricots. Taken together, these results suggest that 1-MCP conferred a greater resistance to oxidative stress. This, along with the reduction in ethylene production, could contribute to the increase in commercial life and nutritional value observed in 1-MCP-treated apricots.


Pilot study of Biomarkers for predicting effectiveness of ramosetron in diarrhea-predominant irritable bowel syndrome: expression of S100A10 and polymorphisms of TPH1.
Serotonin type 3 receptor (5-HT3 R) antagonists are potentially useful therapeutic agents for diarrhea-predominant irritable bowel syndrome (IBS-D). To identify biomarkers predicting effectiveness of the 5-HT3 R antagonist (ramosetron) in IBS-D. Irritable bowel syndrome-D Japanese subjects received 2.5 or 5 μg of ramosetron once daily for 4 weeks. Colonic mucosal S100A and tryptophan hydroxylase (TPH) mRNA expression levels were measured before treatment. Genomic DNA was extracted from blood and polymorphisms of TPH1 and TPH2 were analyzed. Forty-two patients (27 men and 15 women, mean age 42 years) with IBS-D were included for analysis. Improvement of IBS symptoms was seen in 26 (61.9%). Baseline S100A10 (p = 0.02) and TPH1 (p = 0.02) expression were significantly higher in the ramosetron responders than in the non-responders. The frequencies of the TPH1 rs4537731G allele in linkage disequilibrium with the TPH1 rs7130929 T allele (11.5% vs 50%, p = 0.003; OR: 12; 95% CI: 2.1-69) along with TPH1 rs211105 C allele (3.8% vs 43.8%, p = 0.0003; OR: 19; 95% CI: 2.1-181) were significantly lower in the responders than in the non-responders. The mean scores of diarrhea at baseline were significantly higher (5.2 vs 3.7, p = 0.005) in patients with TPH1 rs211105 T/T than those with the G allele. TPH1 gene polymorphisms and S100A10 expression, which correlate with 5-HT signaling were associated with ramosetron effectiveness in IBS-D, and may possibly lead to prospective identification of the resistance to treatment.


Modified culture protocol for isolation of Mycobacterium avium subsp. paratuberculosis from raw milk.
A modified culture method using C18-carboxypropylbetaine (CB-18) and microscopic screening was evaluated for time to and limit of detection of Mycobacterium avium subsp. paratuberculosis (MAP) in raw milk. Bulk-tank milk samples were spiked with six different concentrations (10(1) to 10(6) CFU/mL) of MAP. Samples were processed using two different protocols. The first protocol involved specimen processing with the zwitterionic detergent C18-carboxypropylbetaine (CB-18) and lytic enzymes followed by culture on modified Middlebrook 7H10 agar plates with microscopic screening. The second protocol used 0.75% hexadecylpyridinium chloride (HPC) for specimen processing, followed by culture on Herrold's egg yolk medium (HEYM). Both protocols were repeated eight independent times, and the detection limit and time to detection were compared. The presence of MAP in spiked milk samples was detected between 14 and 45 days (N [number of samples], 46; mean, 22.7; median, 19.5) using the CB-18 and microscopic screening method, and between 21 and 63 days (N, 47; mean, 31; median, 28) using HEYM (P < 0.001). Time to detection also varied with MAP concentration (P < 0.001). Higher concentrations of MAP were detected earlier than lower concentrations and this finding was independent of the method used (P = 0.479). The two methods had similar detection limits but the modified culture method reduced the time to detect MAP in raw milk for the majority of concentrations.


Cholesterol and multilamellar bodies: Lysosomal dysfunction in GBA-Parkinson disease.
Lipid and cholesterol metabolism might play a role in the pathogenesis of Parkinson disease (PD). However, the association between cholesterol and PD is not clearly established. Cholesterol accumulation is closely related to the expression of multilamellar bodies (MLBs). Also, cholesterol controls autophagosome transport. Thus, impaired cholesterol and autophagosome trafficking might lead to robust autophagic vacuole accumulation. Our recent work provides the first evidence that the presence of the N370S GBA mutation produces an accumulation of cholesterol, which alters autophagy-lysosome function with the appearance of MLBs, rendering the cell more vulnerable and sensitive to apoptosis.


[Heterozygous family hypercholesterolemia: study of use of statins under clinical practice conditions].
To compare the use profile of statins in patients with clinical diagnosis of heterozygous family hypercholesterolaemia (HFH) and the use profile of the general population treated with statins. DESIGN. Retrospective, observational study. Galician Health Service, Spain. Patients under 65 years old treated with statins. In patients with HFH: number of patients, age, gender, statins used and defined daily dose per patient and day (DDD/patient/day). Variables in the rest of the population treated with statins: number of patients, statins used and DDD/patient/day. In a sample of 331 under-65 patients with HFH, 185 were men (55.86%) and 146 women (44.14%). Their average age was 44.56 years old (95% CI, 43.33-45.80), without statistically significant differences between men and women. Relevant differences in the daily mean consumption (DDD/patient/day) of statins between the studied sample and the rest of the under-65 population treated with statins were found (3.03 DDD/patient/day [95% CI, 2.70-3.36] vs 1.33 DDD/patient/day [95% CI, 1.16-1.40]; P< .001). These differences were caused by atorvastatin: 70% of HFH patients were being treated with atorvastatin (versus 37.6% of the rest of the population treated with statins). Differences in daily mean consumption of atorvastatin between HFH and the rest of the statin-treated population were also found (3.58 DDD/patient/day [95% CI, 3.15-4.02] vs 1.64 DDD/patient/day [95% CI, 1.37-1.90]; P< .001). The DDD/patient/day of statins in HFH patients is double that of the rest of the population treated with statins. The different pattern of use of atorvastatin in HFH is the main reason for these results.


Detection of sperm-reactive antibodies in wild sika deer and identification of the sperm antigens.
Antisperm antibodies potentially inhibit sperm functions causing the sterility in humans and experimentally treated animals. However, there is no information about antisperm antibodies emerging spontaneously in wildlife. In this study, we searched for the sperm-reactive antibodies, spontaneously produced in wild sika deer (Cervus nippon), and identified the sperm antigens. We collected 529 fecal masses of sika deer in Japanese cities, from which we extracted the mucosal antibodies to test them for reactivities to deer sperm proteins by ELISA. Two of the extracts contained IgAs that were highly reactive to the sperm proteins. The molecular weights of the active IgAs, partially purified by DEAE-sephadex A-50, were estimated at more than 100 kDa, suggesting that the IgAs evaded drastic digestion in the gastrointestinal tract. Two-dimensional electrophoresis and immunoblotting detected three major antigens, and the following LC-MS/MS analysis identified them as alpha-enolase, phosphoglycerate kinase 2 and acrosin-binding protein. The antibodies were cross-reactive to a recombinant human acrosin-binding protein. To our knowledge, this is the first research to find that the sperm-reactive antibodies are produced spontaneously in wildlife and they recognize a common antigen found in humans.


Expression of decidual prolactin-related protein in the rat decidua.
It is well established that rat decidual tissue produces a PRL-like hormone(s) that binds to the PRL receptor on both the corpus luteum and the decidual cells and initiates profound changes in the endocrine milieu required for the establishment of pregnancy. The recent cloning of a decidual PRL-related protein (dPRP) prompted us 1) to determine whether the expression of this gene is triggered by decidualization of the endometrial stromal cells, 2) to examine the temporal and cell-specific pattern of its expression, and 3) to examine the role of both decidual signals and PRL on levels of its messenger RNA (mRNA). Total RNA was isolated from uteri of either nonpseudopregnant rats or pseudopregnant rats with or without decidual tissue. A 1-kilobase mRNA species hybridizing strongly with the dPRP probe was present in decidualized uteri. No dPRP mRNA could be detected in uteri not subjected to decidualization. Developmental studies indicated a constant high level of dPRP mRNA in the decidual tissue until day 12 of pseudopregnancy, followed by a marked decline at a time when extensive cell death occurs in the decidua, suggesting that dPRP is constitutively expressed in this tissue. To examine the cell-specific expression of dPRP, antimesometrial decidua was separated from mesometrial decidua, and the large antimesometrial cell population was separated from the small mesometrial cells by elutriation. The results of Northern analysis revealed clearly that dPRP is abundantly and solely expressed in the large antimesometrial cells. No dPRP mRNA could be detected in the mesometrial cells and in numerous other endocrine and nonendocrine tissues. A faint signal was observed, however, in the trophoblast. Despite the very strong paracrine regulation between the antimesometrial and mesometrial cells and the high levels of PRL receptor expression in these cells, both in vivo and coculture experiments revealed no regulation of dPRP gene expression by either PRL or mesometrial cell signal, adding further support to the possibility that once induced, dPRP remains constitutively expressed. In summary, the results of this investigation revealed that the expression of dPRP in endometrial stromal cells is triggered by the induction of decidualization and that this gene is selectively and abundantly expressed in a defined cell population located in the anti-mesometrial region of the uterus. Thus, dPRP is not only a useful indicator of decidualization, but is also an excellent marker for the differentiated antimesometrial cells.


Reduced grid-cell-like representations in adults at genetic risk for Alzheimer's disease.
Alzheimer's disease (AD) manifests with memory loss and spatial disorientation. AD pathology starts in the entorhinal cortex, making it likely that local neural correlates of spatial navigation, particularly grid cells, are impaired. Grid-cell-like representations in humans can be measured using functional magnetic resonance imaging. We found that young adults at genetic risk for AD (APOE-ε4 carriers) exhibit reduced grid-cell-like representations and altered navigational behavior in a virtual arena. Both changes were associated with impaired spatial memory performance. Reduced grid-cell-like representations were also related to increased hippocampal activity, potentially reflecting compensatory mechanisms that prevent overt spatial memory impairment in APOE-ε4 carriers. Our results provide evidence of behaviorally relevant entorhinal dysfunction in humans at genetic risk for AD, decades before potential disease onset.


Amplitude death in nonlinear oscillators with nonlinear coupling.
Amplitude death is the cessation of oscillations that occurs in coupled nonlinear systems when fixed points are stabilized as a consequence of the interaction. We show here that this phenomenon is very general: it occurs in nonlinearly coupled systems in the absence of parameter mismatch or time delay although time-delayed interactions can enhance the effect. Application is made to synaptically coupled model neurons, nonlinearly coupled Rössler oscillators, as well as to networks of nonlinear oscillators with nonlinear coupling. By suitably designing the nonlinear coupling, arbitrary steady states can be stabilized.


Clinical evaluation of recombinant factor IX.
The pharmacokinetics, safety, and efficacy of recombinant factor IX (rFIX) have been evaluated in previously treated and untreated patients with hemophilia B. In a study of 56 previously treated patients, there was a total of 1,070 hemorrhages, which required 1,514 infusions of rFIX. Of the 1,070 episodes, 80% resolved after a single rFIX infusion. The majority of the remaining episodes requiring more than 1 infusion were either major or complicated hemorrhages. Eighty-seven percent of the 1,514 infusions were rated by the physicians or patients as providing an excellent or good response. These results are consistent with the efficacy data from other hemophilia product clinical trials. rFIX has also been administered as bolus doses or by continuous infusion in conjunction with 13 surgical procedures. Estimated blood loss during and after surgery was as expected when compared with blood loss for similar procedures in patients without hemophilia, and 97% of the clinical responses during and after surgery were rated by physicians or patients as excellent or good. Pharmacokinetic comparison of rFIX with one plasma-derived product showed a statistically significant lower recovery (28% less) of rFIX. The half-lives between the two products were comparable. Common adverse events associated with rFIX treatment have been comparable to other factor IX products. As expected, there has been no evidence of viral transmission by rFIX.


Resistance exercise increases active MMP and β1-integrin protein expression in skeletal muscle.
Recent studies indicate that matrix metalloproteinases (MMPs) and critical linkage proteins in the extracellular matrix (ECM) regulate skeletal muscle mass, although the effects of resistance training (RT) on protein expression and activity are unclear. Thus, the purpose of the present study was to investigate the effects of RT on MMP activity and expression of ECM-related proteins. Ten male Sprague-Dawley rats were randomly assigned to 1 bout (1B) or 18 bouts (18B) of electrical stimulation. The right gastrocnemius muscle was isometrically contracted via percutaneous electrical stimulation (five sets of 5 sec stimulation × five contractions/set with 5 sec interval between contractions and 3 min rest between sets) once (1B) or every other day for 5 weeks (18B). The left leg served as a control. Activity of MMP-2 and MMP-9, determined via gelatin zymography, was increased (P < 0.05) immediately after 1B. However, MMP activation was not evident following 18B. No changes in collagen IV, laminin α2, α7-integrin, or ILK protein expression were detected immediately following 1B or 18B. However, β1-integrin protein expression was significantly increased (P < 0.05) with 18B. Our results suggest that resistance exercise activates MMPs during the initial phase of RT but this response is attenuated with continuation of RT.


Exposure to Stress Alters Cardiac Gene Expression and Exacerbates Myocardial Ischemic Injury in the Female Murine Heart.
Mental stress is a risk factor for myocardial infarction in women. The central hypothesis of this study is that restraint stress induces sex-specific changes in gene expression in the heart, which leads to an intensified response to ischemia/reperfusion injury due to the development of a pro-oxidative environment in female hearts. We challenged male and female C57BL/6 mice in a restraint stress model to mimic the effects of mental stress. Exposure to restraint stress led to sex differences in the expression of genes involved in cardiac hypertrophy, inflammation, and iron-dependent cell death (ferroptosis). Among those genes, we identified tumor protein p53 and cyclin-dependent kinase inhibitor 1A (p21), which have established controversial roles in ferroptosis. The exacerbated response to I/R injury in restraint-stressed females correlated with downregulation of p53 and nuclear factor erythroid 2-related factor 2 (Nrf2, a master regulator of the antioxidant response system-ARE). S-female hearts also showed increased superoxide levels, lipid peroxidation, and prostaglandin-endoperoxide synthase 2 (Ptgs2) expression (a hallmark of ferroptosis) compared with those of their male counterparts. Our study is the first to test the sex-specific impact of restraint stress on the heart in the setting of I/R and its outcome.


Stimulation of enzyme synthesis by sublethal concentrations of chloramphenicol is not mediated by ribonucleotide pools.
Growth of Salmonella typhimurium LT-2 strain pyrA81 in minimal medium containing 0.8 mug of chloramphenicol (CAP) per ml resulted in a 50 to 100% increase in the steady-state nucleotide pools in the cells. When such a culture was starved for uracil, the pyrimidine nucleotide pools decayed much more slowly in CAP-treated cells than in controls. An attempt was made to determine whether this effect of CAP on nucleotide pools could account for enhanced enzyme derepression observed under the same conditions (8). Treatment with low levels of puromycin also resulted in elevation of nucleotide pools but did not lead to enhanced enzyme synthesis. CAP treatment during arginine starvation has been shown to enhance enzyme synthesis, but nucleotide pools were not significantly affected by CAP nor was the stringent response relieved under these conditions. Thus, the effects of CAP on enzyme synthesis cannot be the result of effects of the antibiotic on nucleotide pools in all cases. The elevation of nucleotide pools was shown not to be due to enhanced conversion of exogeneous uracil into nucleotides, but to result from the enhanced turnover of ribonucleic acid in CAP-treated cells.


[Study on dental cement. Part 4. Protective effect of a newly designed light-cured varnish on the vulnerability to water of glass polyalkenoate cement restorations].
The protective effect of a newly designed light-cured varnish on the vulnerability to water of two kinds of glass polyalkenoate cement restorations was examined by determining the penetration depth of Ag ion into the set cement on the SEM pictures in comparison with those of the conventional varnish and a light-cured resin bonding agent. The protective effect of the experimental light-cured varnish was perfect, while those of the conventional varnish and the light-cured resin bonding agent were inferior to that.


[Structure and function of mammalian brain microtubule-associated proteins].
Cytoplasmic microtubules are fibrous intracellular organelles found in almost all eukaryotic cells and play an important role in maintenance of cell shape, cell division, axonal transport, secretion and receptor activity. Besides tubulin dimers, microtubule proteins consist of several other components called MAPs which promote microtubule assembly and form long filamentous projection on the surface of the polymer. In mammalian brain, two classes of MAPs have been characterized; one is structural MAPs including MAP1 (1A and 1B), MAP2 (2A, 2B and 2C) and tau which function in the morphogenesis and maintenance of neural tissues and cells, and the other contains motor MAPs (kinesin and MAP1C) which are related to translocation of vesicles along microtubules in axon and to mitosis. The primary sequences of MAPs have been determined from their cDNAs. The functions of structural MAPs are modulated by their binding to other intracellular components, different expressions of isoforms during brain development and phosphorylation-dephosphorylation by various protein kinases and phosphatases. Biochemical characterization of MAP2 and tau have been well investigated. However, little is known about the function of MAP1 under the biochemical level, because MAP1 is unstable and high sensitive to proteases. We have developed a simple and rapid purification procedure for MAP1 using poly (L-aspartic acid) and taxol, and observed MAP1-F-actin interaction as well as MAP1-microtubules interaction. Recently, we have found that three specific kinases which can phosphorylate MAP1A and 1B are associated with MAP1 preparation and called it MAP1 kinase. Some evidence suggest that one of them is an unknown kinase and others are casein kinase I- and II-like kinases. Further studies to examine MAP1 kinase and phosphorylation of MAP1 provide a valuable insight for understanding thoroughly the microtubule-mediated functions.


New approach for the diagnosis of histamine intolerance based on the determination of histamine and methylhistamine in urine.
Histamine intolerance is a disorder in the homeostasis of histamine due to a reduced intestinal degradation of this amine, mainly caused by diamine oxidase (DAO) enzyme deficiency, which provokes its accumulation in plasma and the appearance of adverse health affects. A new approach for the diagnosis of this intolerance could be the determination of histamine and its metabolites in urine. The aim of this work was to develop and validate a rapid method to determine histamine and methylhistamine in human urine by Ultra High Performance Liquid Chromatography and Fluorimetric detection (UHPLC-FL). The proposed method is a consistent procedure to determine histamine and methylhistamine in less than 11min with adequate linearity and sensitivity. Relative standard deviation was always lower than 5.5%, ensuring method precision; and mean recovery was greater than 99% for both analytes. The structure of histamine and methylhistamine conjugated with OPA were confirmed by UHPLC-ITD-FTMS which enabled to unequivocally identify both analytes in standards and also in urine samples. The analysis of histamine and methylhistamine in urine samples could be a potential new approach for the routine diagnosis of histamine intolerance, more patient-friendly and with clear advantages in terms of equipment and personnel demand for sample collection in comparison with current plasmatic DAO activity determination.


Characterization of two glutaminases from the filamentous cyanobacterium Anabaena sp. PCC 7120.
The Anabaena genome contains two ORFs that appear to encode glutaminases. The genes were expressed as histidine-tagged fusion proteins in Escherichia coli. The purified proteins possessed glutaminase activity using l-glutamine as the substrate, but differed in biochemical properties. All2934 showed an optimal activity at 20 degrees C and pH 6.0, with a higher affinity for l-glutamine than All4774, which had optimal activity at 37 degrees C and pH 7.5. Remarkably, the glutaminase activity of All2934 was phosphate dependent, while All4774 was phosphate independent. The expression of all2934 and all4774 was analyzed using semi-quantitative reverse transcriptase-PCR. The expression level of all2934 was much higher than that of all4774 under normal and nitrogen-depletion conditions, indicating that All2934 may play an important role in metabolizing glutamine in Anabaena.


"On-off" switchable bioelectrocatalysis synergistically controlled by temperature and sodium sulfate concentration based on poly(N-isopropylacrylamide) films.
In this work, poly(N-isopropylacrylamide) (PNIPAm) films were synthesized on a Au electrode surface through the electrochemically induced free-radical polymerization method. The "coil-to-globule" phase transition of PNIPAm films was sensitive to both environmental temperature and sodium sulfate (Na(2)SO(4)) concentration in solution and was detected by cyclic voltammetry (CV) of ferrocenecarboxylic acid (Fc(COOH)) probe. For example, in solutions containing no Na(2)SO(4) at 25 degrees C, the probe demonstrated a well-defined CV peak pair with large peak currents, showing the "on" state; at 35 degrees C, the CV response was significantly suppressed, showing the "off" state. By switching the film electrodes in solution between 25 and 35 degrees C, the CV peak currents cycled between the on and off states, demonstrating the reversible thermosensitive switching function of the films. Similarly, the reversible Na(2)SO(4)-concentration-sensitive "on-off" property of PNIPAm films toward Fc(COOH) was also observed. In particular, the influence of temperature and Na(2)SO(4) concentration on the on-off behavior of the films was not independent or separate, but synergetic or cooperative. The dual-responsive property of the films could also be used to switch the on-off bioelectrocatalysis. That is, the electrochemical oxidation of glucose catalyzed by glucose oxidase (GOx) and mediated by Fc(COOH) in solution could be controlled or modulated by changing the surrounding temperature, Na(2)SO(4) concentration, or both. This dual- and synergetic-triggered bioelectrocatalysis based on the "smart" PNIPAm interface system may establish a foundation for fabricating novel multiple factor-controllable biosensors.


Transplantation of fibroblasts that overexpress matrix metalloproteinase-3 into the site of spinal cord injury in rats.
Growth-inhibitory chondroitin sulfate proteoglycans (CSPGs) are upregulated during glial scar formation at the site of spinal cord injury (SCI) in adult mammals. This CSPG-containing glial scar inhibits axonal regeneration. Matrix metalloproteinases (MMPs) can degrade CSPGs and other inhibitory proteins to promote neurite outgrowth. Increased MMP synthesis and secretion are observed in fibroblasts adjacent to tumor cells that express the protein EMMPRIN (Extracellular Matrix MetalloPRoteinase INducer). EMMPRIN transduction of cells provides an avenue to deliver increased levels of MMPs to the site of SCI in a sustained, localized, and moderate fashion. We explored the use of EMMPRIN-transduced cells as a mechanism to degrade CSPGs, facilitate axonal growth and improve recovery after SCI. Human dermal fibroblasts infected with a recombinant EMMPRIN adenovirus significantly increased secretion of MMP-3 compared to fibroblasts infected with a control adenovirus. Decreased CSPG immunoreactivity was observed in injured spinal cord sections when they were incubated with media from EMMPRIN-transduced fibroblasts. Conditioned media from EMMPRIN-transduced fibroblasts increased the length of neurites that were grown on a CSPG substrate. Rats that received contusive SCI and EMMPRIN-transduced fibroblast transplants demonstrated improved locomotor recovery compared to rats that received control fibroblasts, but not compared to other control groups. EMMPRIN-transplanted rats showed a significant increase in the number of retrogradely labeled cell bodies within brainstem nuclei and an increase in serotonergic fibers distal to the site of injury. EMMPRIN, and consequently MMP, delivery to the injured spinal cord may prove to be beneficial in reducing some of the physical barriers to axonal growth after SCI.


High yields of influenza A virus in Madin-Darby canine kidney cells are promoted by an insufficient interferon-induced antiviral state.
Because of their high susceptibility to infection with various influenza virus strains, Madin-Darby canine kidney (MDCK) cells have been widely used as a substrate for influenza virus isolation and vaccine production. However, MDCK cells are also interferon (IFN) competent, and the type I IFN response is commonly thought to be a factor strongly inhibiting virus replication. Therefore, the inhibition of influenza virus replication by IFN signalling was analysed for an adherent MDCK cell line used in vaccine manufacturing. Depending on the respective virus strain, different levels of IFN induction and a corresponding upregulation of the IFN-induced myxovirus resistance protein 1 (Mx1) were observed. Suppression of IFN induction by transient expression of the viral non-structural protein 1 protein enhanced replication of an influenza virus lacking NS1, but not wild-type strains. In agreement with this, stimulation of cells with MDCK cell-derived IFN prior to infection resulted only in a decrease in replication rate, and not in a change of final yields for wild-type influenza viruses. This lack of IFN-induced antiviral activity correlated with missing anti-influenza activity of MDCK Mx proteins. No inhibitory effect on viral polymerase activity was found for canine Mx1 (cMx1) and cMx2 in minireplicon assays. In conclusion, in MDCK cells, IFN expression is not a limiting factor for influenza virus replication and this might partially be caused by a lack of anti-influenza activity of canine Mx proteins.


Role of the mitochondrial permeability transition in apoptotic and necrotic death after ischemia/reperfusion injury to hepatocytes.
Reperfusion of ATP-depleted tissues after warm or cold ischemia causes pH-dependent necrotic and apoptotic cell death. In hepatocytes and other cell types as well, the mechanism underlying this reperfusion-induced cell death involves onset of the mitochondrial permeability transition (MPT). Opening of permeability transition (PT) pores in the mitochondrial inner membrane initiates the MPT, an event blocked by cyclosporin A (CsA) and pH less than 7.4. Thus, both acidotic pH and CsA prevent MPT-dependent reperfusion injury. Glycine also blocks reperfusion-induced necrosis but acts downstream of PT pore opening by stabilizing the plasma membrane. After the MPT, ATP availability from glycolysis or other source determines whether cell injury after reperfusion progresses to ATP depletion-dependent necrosis or ATP-requiring apoptosis. Thus, apoptosis and necrosis after reperfusion share a common pathway, the MPT. Cell injury progressing to either necrosis or apoptosis by shared pathways can be more aptly termed necrapoptosis.


Determinants of father involvement with young children: Evidence from the Early Childhood Longitudinal Study-Birth Cohort.
The current study used data from the Early Childhood Longitudinal Study-Birth Cohort (ECLS-B; Snow et al., 2007) to explore determinants of resident father involvement. Families (N = 2,900) were measured at 3 time points (9 months, 2 years, and 4 years of age). Father, mother, and child factors were examined in relation to father caregiving and play. Latent change score models indicated that fathers engaged in more caregiving and play behaviors and increased at a faster rate when they more strongly identified with their role as a father. Fathers engaged in more caregiving when mothers reported higher depressive symptoms and increased in play more slowly when marital conflict was higher. In addition, a Mother Depressive Symptoms × Marital Conflict interaction emerged indicating that fathers differed in their levels of caregiving depending on mothers' report of depressive symptoms, but only when marital conflict was low. Fathers also increased in caregiving at a faster rate with girls than boys. A comprehensive framework for examining resident father involvement is presented.


Bovine enzootic haematuria from consumption of Pteris deflexa and Pteris plumula in northwestern Argentina.
Bovine enzootic haematuria (BEH) is caused by prolonged ingestion of toxic principles of bracken fern, essentially by Pteridium spp. In northwestern Argentina, this disease has a great economic impact ant it is attributed a chronic consumption to Pteridium arachnoideum. This paper describes two endemic areas for enzootic hematuria due to the consumption of Pteris deflexa and Pteris plumula. Two areas where P. deflexa and P. plumula are endemic, but free of Pteridium species, were devised and seven farms were visited. The disease was confirmed based on the presence of clinically affected animals. In four necropsies bleeding neoplastic lesions were observed in the mucosa of the urinary bladder. At phytochemical analysis, both ptaquiloside and pterosin B were found in P. deflexa and P. plumula. Thus, the consumption of P. deflexa and P. plumula can also cause BEH.


Examining the Relationship Between Symptomatic Burden and Self-reported Productivity Losses Among Patients With Uterine Fibroids in the United States.
To evaluate the impact of uterine fibroid symptoms on employment and household productivity. An online survey of US women between 18 and 54 was conducted. Productivity was assessed using the health related productivity questionnaire (HRPQ). Descriptive statistics and logistic multivariable regressions examined the relationship between uterine fibroids (UF) symptom experience and employment and household productivity. Of 1365 eligible women, 873 (64.0%) were employed. Women lost an average of 0.8 hours to employment-related absenteeism and 4.4 hours due to employment-related presenteeism for 5.1 hours of employment productivity lost/week. Women lost an average of 1.4 hours due to household-related absenteeism and 1.6 hours due to household-related presenteeism for a total of 3.0 hours of household lost productivity. Productivity losses increased with increases in symptom burden. UF has a substantial impact on employment-related and household-related productivity.


Dm-myb mutant lethality in Drosophila is dependent upon mip130: positive and negative regulation of DNA replication.
Gene amplification at the chorion loci in Drosophila ovarian follicle cells is a model for the developmental regulation of DNA replication. Previously, we showed that the Drosophila homolog of the Myb oncoprotein family (DmMyb) is tightly associated with four additional proteins and that DmMyb is required for this replication-mediated amplification. Here we used targeted mutagenesis to generate a mutant in the largest subunit of the DmMyb complex, the Aly and Lin-9 family member, Myb-interacting protein 130 (Mip130). We found that mip130 mutant females are sterile and display inappropriate bromodeoxyuridine (BrdU) incorporation throughout the follicle cell nuclei at stages undergoing gene amplification. Whereas mutations in Dm-myb are lethal, mutations in mip130 are viable. Surprisingly, Dm-myb mip130 double mutants are also viable and display the same phenotypes as mip130 mutants alone. This suggests that Mip130 activity without DmMyb counteraction may be responsible for the Dm-myb mutant lethality. RNA interference (RNAi) to selectively remove each DmMyb complex member revealed that DmMyb protein levels are dependent upon the presence of several of the complex members. Together, these data support a model in which DmMyb activates a repressive complex containing Mip130 into a complex competent to support replication at specific loci in a temporally and developmentally proscribed manner.


Staphylococcal superantigen-like protein 3 binds to the Toll-like receptor 2 extracellular domain and inhibits cytokine production induced by Staphylococcus aureus, cell wall component, or lipopeptides in murine macrophages.
Staphylococcal superantigen-like proteins (SSLs) are a family of exoproteins sharing structural similarity with superantigens, but no superantigenic activity. Corresponding host target proteins or receptors against a portion of SSLs in the family have been identified. In this study, we show that SSL3 specifically binds to Toll-like receptor 2 (TLR2) and inhibits the stimulation of macrophages by TLR2 ligands. An approximately 100-kDa protein was recovered by using recombinant His-tagged SSL3-conjugated Sepharose from the lysate of porcine spleen, and the protein was identified as porcine TLR2 by peptide mass fingerprinting analysis. The SSL3-conjugated Sepharose recovered human and mouse TLR2 but not TLR4 from human neutrophils and mouse macrophage RAW 264.7 cells, as well as a recombinant TLR2 extracellular domain chimera protein. The production levels of interleukin 12 (IL-12) from mouse macrophages treated with heat-killed Staphylococcus aureus and of tumor necrosis factor alpha (TNF-α) from RAW 264.7 cells induced by peptidoglycan or lipopeptide TLR2 ligands were strongly suppressed in the presence of SSL3. The mutation of consensus sialic acid-containing glycan-binding residues in SSL3 did not abrogate the binding ability to TLR2 or inhibitory activity on TLR2, indicating that the interaction of SSL3 with TLR2 was independent of the sialic acid-containing glycan-binding residues. These findings demonstrate that SSL3 is able to bind the extracellular domain of TLR2 and interfere with TLR2 function. The present study provides a novel mechanism of SSL3 in immune evasion of S. aureus via interfering with its recognition by innate immune cells.


Renaissance of the biologically active vitamin A derivatives: established and novel directed therapies for cancer and chemoprevention.
Vitamin A and its biologically active derivatives are involved in a complex arrangements of physiological and developmental responses in many tissue of higher vertebrates. Retinoids are natural and synthetic compounds related to retinoic acid that act through interaction with two basic types of nuclear receptors: retinoic acid receptors (RAR alpha, RARbeta and RARgamma) and retinoid X receptors (RXRalpha, RXRbeta and RXRgamma) as retinoid-inducible transcription factors. Thus, the retinoid receptors are considered to be ligand-activated, DNA-binding, trans-acting, transcription-modulating proteins involved in a general molecular mechanism responsible for transcriptional responses in target genes. They exert both beneficial and detrimental activity; they have tumor-suppressive activity but on the other hand they are teratogenic. Retinoids inhibit carcinogenesis, suppress premalignant epithelial lesions and tumor growth and invasion in a variety of tissues. Natural and synthetic retinoids have therapeutical effects due to their antiproliferative and apoptosis-inducing effects. They are known to cause redifferentiation or to prevent further dedifferentiation of various neoplastic tissues. A number of novel chemical compounds, receptor selective retinoids and rexinoids, have been synthesized up to now and tested both in vitro and in vivo, using animal models against different cancer cells. In spite of that progress, there is still an urgent call for novel synthetic retinoids and rexinoids with greater retinoid receptor selectivity, reasonable chemotherapeutic or chemopreventive effects and reduced toxicity and side effects. This article summarizes selected effects of biologically active natural or synthetic retinoids and rexinoids, acting through their cognate nuclear receptors, and their use in chemotherapy and chemoprevention of various types of cancer.


Olfactory vector hypothesis for encephalitis lethargica.
Viruses have long been implicated in the pathogenesis of classical encephalitis lethargica, which was first described by Constantin von Economo in 1917. In this article, I propose the hypothesis that an airborne virus travels along the olfactory conduit to infect the olfactory bulb; this local infection or induced neuroinflammation, in turn, retrogradely targets certain neuronal populations with sleep-wake regulatory functions in the hypothalamus and midbrain, leading to the development of wakeful inactivity, a hallmark clinical feature of the disease. Furthermore, the olfactory vector hypothesis may also explain the pathomechanism of the debilitating complication of the disease, i.e., postencephalitic parkinsonism, in terms of a recently discovered nigro-olfactory projection.


Protective effect of MAAs extracted from Porphyra tenera against UV irradiation-induced photoaging in mouse skin.
The objective of this study was to investigate the effect of Mycosporine-like amino acids (MAAs) extracted from Porphyra tenera skin against UV irradiation-induced photoaging using an ICR mouse model of skin photoaging and to explore the curative effects of the compounds in MAAs. The skin damage and collagenous tissue impairments induced by ultraviolet radiation were observed by histopathological analysis, and the effects of MAAs on protecting against skin damage and maintaining an intact structure of collagenous tissue were studied. The expression of NF-κB and the MAPK signaling pathway and nuclear transcription factors MMP-1, MMP-3 and TNF-α was analyzed used quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) and ELISA assays. The results showed that the MAAs extracted from Porphyra tenera contained Porphyra-334 and shinorine, which could prevent skin photoaging induced by ultraviolet irradiation and reduce the damage to collagen and elastin. Meanwhile, MAAs significantly inhibited the decrease in hydroxyproline and collagen content and protected against pathological damage to collagen fibers in photoaging skin. MAAs resulted in a reduction in the expression of interstitial collagenase (MMP-1), matrix metalloproteinases (MMP-3) and tumor necrosis factor-α (TNF-α) and a reduction in the content of tissue matrix metalloproteinase (MMPs). Furthermore, MAAs may promote procollagen synthesis by downregulating the expression of TNF-α and downregulating the levels of MMPs, which demonstrates that MAAs are involved in matrix collagen synthesis by activating the NF-kB pathway in photoaging skin. Based on these results, we concluded that MAAs protect skin from UV irradiation-induced photodamage, and therefore, MAAs may be a potentially effective agent for the prevention of photoaging.


Multiple-relaxation-time lattice Boltzmann models in three dimensions.
This article provides a concise exposition of the multiple-relaxation-time lattice Boltzmann equation, with examples of 15-velocity and 19-velocity models in three dimensions. Simulation of a diagonally lid-driven cavity flow in three dimensions at Re = 500 and 2000 is performed. The results clearly demonstrate the superior numerical stability of the multiple-relaxation-time lattice Boltzmann equation over the popular lattice Bhatnagar-Gross-Krook equation.


Geometric Response and Disclination-Induced Skin Effects in Non-Hermitian Systems.
We study the geometric response of three-dimensional non-Hermitian crystalline systems with nontrivial point-gap topology. For systems with fourfold rotation symmetry, we show that in the presence of disclination lines with a total Frank angle, which is an integer multiple of 2π, there can be nontrivial one-dimensional point-gap topology along the direction of the disclination lines. This results in disclination-induced non-Hermitian skin effects. By doubling a non-Hermitian Hamiltonian to a Hermitian three-dimensional chiral topological insulator, we show that the disclination-induced skin modes are zero modes of the effective surface Dirac fermion(s) in the presence of a pseudomagnetic flux induced by disclinations. Furthermore, we find that our results have a field theoretic description, and the corresponding geometric response actions (e.g., the Euclidean Wen-Zee action) enrich the topological field theory of non-Hermitian systems.


An anatomical study of the origins of the medial circumflex femoral artery in the Turkish population.
The medial circumflex femoral artery (MCFA) usually branches from the deep femoral artery (DFA). It may also branch from the femoral artery (FA). In this study 100 inguinal regions of 50 cadavers were investigated. In 79 extremities (79%) MCFA branched from DFA, while in 15 (15%) it branched from FA. In four extremities (4%) MCFA was found to be double; in each case one of MCFAs branched from FA and the other from DFA. In one of these four cases the lateral circumflex femoral artery (LCFA) was also double. In one case we found a common trunk of DFA and MCFA and in another case a common trunk of MCFA, DFA and LCFA. Clinicians must be familiar with the variations of this clinically important artery to improve their success in the diagnosis and treatment of pathologies in the region.


Improvement of water quality with probiotics inclusion during simulated transport of yellowfin seabream (Acanthopagrus latus) larvae.
The expansion of marine aquaculture requires the shipment of early life stages from hatcheries to breeding centers. Thus, maintaining water quality parameters during transportation is crucial. This experiment investigated the effects of a commercial probiotic mixture (PM) (consisting of Bacillus licheniformis, Bacillus subtilis, Pedicoccus acidilactici, and Lactobacillus acidophilus with a total count of 107 CFU/g) on water quality, bacterial population, and the survival of Acanthopagrus latus larvae during a 24-hour simulated transportation experiment. To simulate transportation, purified seawater was used as control and purified seawater supplemented with 3 g of PM was used as the PM48 treatment. For the PM48 treatment, 3 g of PM was added to 1 L of purified seawater and mixed with 200 L of purified seawater 48 hours prior to the main experiment. The use of probiotics resulted in an increase in larval survival up to 93.3% compared to 89.85% in the control group. Initial sampling showed a statistically significant difference in NO2 - levels between the control and PM48 treatments. After transportation, the pH of the control treatment showed a significant decrease. The total number of bacteria and the total number of Vibrio spp. in water and fish larvae showed that within 48 hours, the probiotics became the dominant population in the PM48 treatment. After 24 hours, water samples and larvae from the control treatment showed a significant increase in bacterial load. In conclusion, the results showed that probiotics can play a significant role in maintaining biotic and abiotic water quality parameters, leading to a better survival rate of larvae during transportation. These findings represent an important step in improving long-distance transport protocols.


Reversible cerebral segmental vasoconstriction.
Vasoconstriction is not recognized as a cause of cerebrovascular disease except in the vasospasm seen following subarachnoid hemorrhage and possibly in migraine. However, we found four patients to have transient, fully reversible vasoconstriction and dilatation prominently involving arteries around the circle of Willis. All four patients were evaluated for severe headaches and fluctuating or recurring motor or sensory deficits. No cause for the clinical syndromes and angiographic abnormalities was found. Similar patients are reported in the literature under various nosologies. This newly recognized clinical-angiographic syndrome should be differentiated from other known causes of vessel constriction and dilatation; the precipitants of reversible vasoconstriction may then be better defined.


A multipronged approach to addressing the organ shortage.
Organ donation rates continue to lag behind the number of patients waiting for an organ transplant. The Alabama Organ Center, the organ procurement organization for the state of Alabama, is using a multipronged approach to address the organ shortage. The plan includes clinical options, public education, and professional education. Donation after cardiac death is proving to be an excellent option to increase the number of donors. Public and professional education must be conducted through a variety of avenues, and the Alabama Organ Center has implemented a successful Department of Motor Vehicles educational effort, in conjunction with other programs. The current issue impacting professional education is participation in the Health and Human Services Organ Donation Breakthrough Collaborative. The common link to all of these programs is effective relationship building.


Eps8 controls actin-based motility by capping the barbed ends of actin filaments.
Actin filament barbed-end capping proteins are essential for cell motility, as they regulate the growth of actin filaments to generate propulsive force. One family of capping proteins, whose prototype is gelsolin, shares modular architecture, mechanism of action, and regulation through signalling-dependent mechanisms, such as Ca(2+) or phosphatidylinositol-4,5-phosphate binding. Here we show that proteins of another family, the Eps8 family, also show barbed-end capping activity, which resides in their conserved carboxy-terminal effector domain. The isolated effector domain of Eps8 caps barbed ends with an affinity in the nanomolar range. Conversely, full-length Eps8 is auto-inhibited in vitro, and interaction with the Abi1 protein relieves this inhibition. In vivo, Eps8 is recruited to actin dynamic sites, and its removal impairs actin-based propulsion. Eps8-family proteins do not show any similarity to gelsolin-like proteins. Thus, our results identify a new family of actin cappers, and unveil novel modalities of regulation of capping through protein-protein interactions. One established function of the Eps8-Abi1 complex is to participate in the activation of the small GTPase Rac, suggesting a multifaceted role for this complex in actin dynamics, possibly through the participation in alternative larger complexes.


Measurement of lifespan in Drosophila melanogaster.
Aging is a phenomenon that results in steady physiological deterioration in nearly all organisms in which it has been examined, leading to reduced physical performance and increased risk of disease. Individual aging is manifest at the population level as an increase in age-dependent mortality, which is often measured in the laboratory by observing lifespan in large cohorts of age-matched individuals. Experiments that seek to quantify the extent to which genetic or environmental manipulations impact lifespan in simple model organisms have been remarkably successful for understanding the aspects of aging that are conserved across taxa and for inspiring new strategies for extending lifespan and preventing age-associated disease in mammals. The vinegar fly, Drosophila melanogaster, is an attractive model organism for studying the mechanisms of aging due to its relatively short lifespan, convenient husbandry, and facile genetics. However, demographic measures of aging, including age-specific survival and mortality, are extraordinarily susceptible to even minor variations in experimental design and environment, and the maintenance of strict laboratory practices for the duration of aging experiments is required. These considerations, together with the need to practice careful control of genetic background, are essential for generating robust measurements. Indeed, there are many notable controversies surrounding inference from longevity experiments in yeast, worms, flies and mice that have been traced to environmental or genetic artifacts(1-4). In this protocol, we describe a set of procedures that have been optimized over many years of measuring longevity in Drosophila using laboratory vials. We also describe the use of the dLife software, which was developed by our laboratory and is available for download (http://sitemaker.umich.edu/pletcherlab/software). dLife accelerates throughput and promotes good practices by incorporating optimal experimental design, simplifying fly handling and data collection, and standardizing data analysis. We will also discuss the many potential pitfalls in the design, collection, and interpretation of lifespan data, and we provide steps to avoid these dangers.


Creatine phosphokinase after submaximal physical exercise in untrained individuals.
Serial estimations of total serum creatine phosphokinase (CPK) have been performed before and during 18-49 hours after submaximal physical exercise in 17 untrained individuals, mean age 50 years. The maximal CPK increase after exercise was 32 mU/ml (73%). The serum CPK did not exceed the upper normal limit (130 mU/ml) except in one individual (150 mU/ml). The maximal CPK increase in patients with acute myocardial infarction (AMI) varied between 101 mU/ml (133%) and 2 260mU/ml(3 790%), mean 900 mU/ml (1 184%). As the maximal CPK elevation in AMI occurs within the same period, it seems that heavy physical work of short duration just before the onset of symptoms will very seldom impair the diagnosis of AMI with the CPK technique used.


Expression of retrovirally transduced genes in primary cultures of adult rat hepatocytes.
Differentiated primary rat hepatocyte cultures have been infected with retroviral vectors expressing human hypoxanthine/guanine phosphoribosyltransferase or the transposon Tn5 neomycin-resistance gene. Expression of the markers was detected only after infection of the cells during a short period of cell replication and transient dedifferentiation from days 1 to 5 of culture. Provirus integrated during that period remains fully expressed during the entire subsequent stationary period of culture up to at least 25 days. Selection with the neomycin analogue G418 of cells infected with the neomycin vector led to the appearance of cells with hepatocyte morphology in which newly synthesized albumin was detectable by immunoprecipitation, indicating successful infection of hepatocytes.


Lethal effect of lambda DNA terminase in recombination deficient Escherichia coli.
lambda DNA terminase is the enzyme that catalyses the cleavage of lambda DNA concatemers into genome-size molecules and packages them into the capsid. The cleavage (DNA maturation) takes place in a specific site in the phage DNA called cos. Either one of two Escherichia coli proteins, integration host factor (IHF) and terminase host factor (THF), is required, in addition to terminase, for maturation of wild-type lambda DNA in vitro. In vivo, at least some cos cleavage is known to occur in mutants that are unable to synthesize active IHF. No THF-defective mutants have yet been isolated. In order to determine if IHF, THF or any other host protein is involved in lambda DNA maturation in vivo, I devised a selection for host mutants that are unable to support cos cleavage. The selection is based on the assumption that lambda DNA terminase will kill cells by cleaving chromosomally located cos sites. I found that DNA terminase will indeed kill cells provided that they contain a chromosomal cos site and provided also that they are defective in the host recA or recB genes. These two genes are required for certain pathways of genetic recombination and repair of damaged DNA, and I suggest that they prevent terminase-induced killing by repairing broken chromosomes. Interestingly, mutation in a related host gene, recD, did not render cells susceptible to terminase killing. recD and recB both encode subunits of exonuclease V, but recD mutants, unlike recB, remain proficient in genetic recombination and repair.(ABSTRACT TRUNCATED AT 250 WORDS)


[Age Related Macular Degeneration--a review].
Age-Related Macular Degeneration is the leading cause of legal blindness in elderly people in Iceland as well as in the western world. If not addressed this will have a huge impact on the quality of life in the ever increasing elderly population. The etiology remains unknown inspite of a better understanding of the pathogenesis. Our treatment options have therefore been limited in the past. For the last few years we have seen a certain progress in drug development and introduction of new drugs into the treatment regime which have reduced severe vision loss somewhat and increased quality of life for these patients. It has also given hope for better treatments to come, which ultimately will be able to stop the disease completely or even prevent it.


Crystal structure of the Legionella pneumophila Lem10 effector reveals a new member of the HD protein superfamily.
Legionella pneumophila, the intracellular pathogen that can cause severe pneumonia known as Legionnaire's disease, translocates close to 300 effectors inside the host cell using Dot/Icm type IVB secretion system. The structure and function for the majority of these effector proteins remains unknown. Here, we present the crystal structure of the L. pneumophila effector Lem10. The structure reveals a multidomain organization with the largest C-terminal domain showing strong structural similarity to the HD protein superfamily representatives. However, Lem10 lacks the catalytic His-Asp residue pair and does not show any in vitro phosphohydrolase enzymatic activity, typical for HD proteins. While the biological function of Lem10 remains elusive, our analysis shows that similar distinct features are shared by a significant number of HD domains found in Legionella proteins, including the SidE family of effectors known to play an important role during infection. Taken together our data point to the presence of a specific group of non-catalytic Legionella HD domains, dubbed LHDs, which are involved in pathogenesis.


Corepressor metastasis-associated protein 3 modulates epithelial-to-mesenchymal transition and metastasis.
Worldwide, metastasis is the leading cause of more than 90% of cancer-related deaths. Currently, no specific therapies effectively impede metastasis. Metastatic processes are controlled by complex regulatory networks and transcriptional hierarchy. Corepressor metastasis-associated protein 3 (MTA3) has been confirmed as a novel component of nucleosome remodeling and histone deacetylation (NuRD). Increasing evidence supports the theory that, in the recruitment of transcription factors, coregulators function as master regulators rather than passive passengers. As a master regulator, MTA3 governs the target selection for NuRD and functions as a transcriptional repressor. MTA3 dysregulation is associated with tumor progression, invasion, and metastasis in various cancers. MTA3 is also a key regulator of E-cadherin expression and epithelial-to-mesenchymal transition. Elucidating the functions of MTA3 might help to find additional therapeutic approaches for targeting components of NuRD.


The ectomycorrhizal association of Tricholoma matsutake and two allied species, T. bakamatsutake and T. fulvocastaneum, with native hosts in subtropical China.
Accurate identification of edible ectomycorrhizal (ECM) mushrooms and their host trees in nature is key to commercial production for consumption. For the first time we describe the ectomycorrhizal association of the three most common species of edible matsutake mushrooms with their native host trees in Yunnan Province in China. We collected ECM samples from three different localities in subtropical forests known to be highly productive areas of Tricholoma. Additionally, we collected basidiomata of Tricholoma from the field and markets in Yunnan. ECM samples were analyzed using morphological and molecular methods. We conducted phylogenetic analyses of nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS) and analyzed the intergenic spacer of cpDNA psbA-trnH to identify basidiomata and plant hosts, respectively. Three species of Tricholoma were identified: T. bakamatsutake, T. fulvocastaneum, and T. matsutake. Four ECM associations in the study area were detected: Tricholoma bakamatsutake + Castanopsis tibetana, T. fulvocastaneum + C. tibetana, T. fulvocastaneum + Pinus yunnanensis, and T. matsutake + P. yunnanensis. Detailed descriptions and illustrations of the ECM associations are presented.


Post-polypectomy electrocoagulation syndrome: a rare cause of acute abdominal pain.
While generally safe, the most feared complication of colonoscopy is perforation of the colon, occurring in nearly 1 in 1,000 procedures, and is more common when polypectomy is performed and electrocautery is used. Less commonly known is the post-polypectomy electrocoagulation syndrome, a transmural burn of the colon which mimics the signs and symptoms of perforation as well as the time course, but follows a benign course and can be treated conservatively.


An isolated unusual digit metastasis from esophageal carcinoma: a case report.
Distant soft tissue tumor metastasis isolated in the digit, presenting as a primary esophageal squamous cell carcinoma is considered extremely rare. Herein, we present a rare case of a 44-year-old male patient with squamous cell carcinoma of the esophagus associated with the clinical symptoms of progressive dysphagia and hoarseness in the course of 2 days. During the second course of chemotherapy, the patient sustained a fall with scald to his right ring finger, while the swelling in the right ring finger was soft, cystic and 2.0×1.8 cm in size. Then, phalangectomy was performed in his right ring finger and pathologic diagnosis was considered metastasis from the esophagus. Unfortunately, the patient succumbed to this disease within 2 months of diagnosis of metastasis. In conclusion, detection of soft tissue metastasis may have prognostic implications, providing more accessible biopsy sites and helping avoid invasive procedures.


A description of a pleurolophocercous cercaria of Procerovum sp from the Haplorchis group of heterophyid trematodes.
Freshwater snails, Stenomelania denisoniensis (Brot) from Tinaroo Dam, North Queensland, Australia were found to be infected with a heterophyid cercaria identified as Procerovum sp. The tail of the cercaria has finfolds which are bilateral anteriorly and dorso-ventral posteriorly, features which separate it from other genera in the Haplorchis-group. This group is differentiated from the cercariae of all the other heterophyid genera by the presence of the penetration glands that extend to the posterior end of the body lateral to the excretory bladder. This paper presents a full description of the cercaria, together with comparisons with other known species of Procerovum.


Selenium-chitosan alleviates the toxic effects of Zearalenone on antioxidant and immune function in mice.
This study assessed the protective effects of selenium-chitosan (SC) against antioxidant and immune function-related damage induced by zearalenone (ZEN) in mice. In total, 150 female mice were allotted to five groups for a 30-day study. Control mice were fed a basal diet. Mice in the ZEN, ZEN-Se1, ZEN-Se2 and ZEN-Se3 groups were fed the basal diet supplemented with same dose of ZEN (2 mg/kg) and different doses of SC, 0.0, 0.2, 0.4 and 0.6 mg/kg, respectively (calculated by selenium). After 30 days, the total antioxidant capacity (T-AOC) level, glutathione peroxidase (GSH-Px) activity, total superoxide dismutase (T-SOD) activity and malondialdehyde (MDA) content in plasma and liver, as well as Con A-induced splenocyte proliferation, plasma interleukins concentrations and liver interleukin mRNA expression levels were determined. The plasma and liver GSH-Px activities, liver T-AOC levels, Con A-induced splenocyte proliferation, interleukin (IL) contents and mRNA expression levels in the ZEN group were significantly lower than in the control group (P < 0.01 or P < 0.05), whereas plasma and liver MDA contents in the ZEN group were significantly higher than in the control group (P < 0.01 or P < 0.05). Additionally, plasma and liver GSH-Px activities, liver T-AOC levels, Con A-induced splenocyte proliferation, IL-1β, IL-17A, IL-2 and IL-6 contents and mRNA expression levels in ZEN+Se2 and ZEN+Se3 groups were significantly higher than in the ZEN group (P < 0.01 or P < 0.05), whereas plasma and liver MDA contents in the ZEN+Se2 and ZEN+Se3 groups were significantly lower than in the ZEN group (P < 0.01 or P < 0.05). The plasma and liver GSH-Px activities, Con A-induced splenocyte proliferation, IL-1β and IL-6 contents, IL-2 and IL-17A mRNA expression levels in the ZEN+Se1 group were also significantly higher than in the ZEN group (P < 0.01 or P < 0.05), whereas the plasma MDA content in the ZEN+Se1 group was also significantly lower than in the ZEN group (P < 0.01). Thus, SC may alleviate antioxidant function-related damage and immunosuppression induced by ZEN in mice.


DNA-inspired oligomers: from oligophosphates to functional materials.
To exert its role of a functional polymer, DNA relies on a molecular self-assembly process that is driven by the interactions of only four units placed in a defined order. Extending the structural diversity of recognition motifs in DNA, to and beyond analogues of the nucleobases, will open doors to self-assembled materials with advanced programmable properties. DNA-inspired systems are becoming useful for numerous applications unachievable by the nucleic acids in their native composition. Potential applications of rationally designed oligo- and polyphosphodiesters reside in the areas of drug delivery, diagnostic signalling and imaging, in systems for efficient energy transfer or the precise ordering on the nanoscale. The field of DNA-inspired phosphodiesters highlights the general value and utility of precision in the composition of oligomers and polymers. In this tutorial review, we will summarize the approaches of directing the self-assembly of DNA-inspired, sequence-specific polyphosphodiesters into soft materials with unique properties. These data expose the so far underexploited potential of DNA-derived systems in solving some of the key issues in various technological areas, such as advanced biomaterials, morphologically defined soft matter or the controlled polymer folding and assembly. Moreover, precise positioning of structurally diverse molecules within a polymer chain creates unmatched opportunities for encoding information on the molecular level and transmitting it further to the microscopic and even macroscopic level via noncovalent interactions.


Perinatal interventions for mothers and fathers who are survivors of childhood sexual abuse.
Childhood sexual abuse (CSA) is a worldwide problem with severe long-term consequences. A history of CSA can impact the childbearing experience of mothers and fathers; affecting their mental health, parenting skills and compromising infant development. Nonetheless, the perinatal period offers huge opportunity for intervention and hope. This literature review collates evidence for perinatal psychosocial interventions targeting both mothers and fathers who are survivors of CSA. Publications dating from 1970 to June 2016 were searched using Medline, Maternity and Infant Health, PsychINFO, PsychArticles, PubMed and the International Bibliography of the Social Sciences (IBSS). There were no perinatal interventions that considered the needs of survivor fathers. Sixteen publications on 9 psychosocial perinatal interventions for CSA survivors were identified. However, no sub-analyses specific to CSA survivors were reported. Trauma-specific perinatal interventions drew from a range of theoretical models and varied widely in format. Generally interventions were associated with improvements in maternal mental health, parenting competence, infant attachment security and positive public health outcomes. They were safe and feasible to implement, acceptable to parents and therapist, and therapists were able to implement protocols with adequate fidelity. Yet current data is hampered by small sample size, inconsistent reporting of CSA rates and outcome measures, scarcity of observational data and longer-term follow-up. Intervention modifications are proposed for CSA survivors in view of their unique childbearing experiences.


Molecular adsorption at electrolyte/α-Al2O3 interface of aluminum electrolytic capacitor revealed by sum frequency vibrational spectroscopy.
The operating voltage of an aluminum electrolytic capacitor is determined by the breakdown voltage (Ub) of the Al2O3 anode. Ub is related to the molecular adsorption at the Al2O3/electrolyte interface. Therefore, we have employed sum-frequency vibrational spectroscopy (SFVS) to study the adsorption states of a simple electrolyte, ethylene glycol (EG) solution with ammonium adipate, on an α-Al2O3 surface. In an acidic electrolyte (pH < 6), the Al2O3 surface is positively charged. The observed SFVS spectra show that long chain molecules poly ethylene glycol and ethylene glycol adipate adopt a "lying" orientation at the interface. In an alkaline electrolyte (pH > 8), the Al2O3 surface is negatively charged and the short chain EG molecules adopt a "tilting" orientation. The Ub results exhibit a much higher value at pH < 6 compared with that at pH > 8. Since the "lying" long chain molecules cover and protect the Al2O3 surface, Ub increases with a decrease of pH. These findings provide new insights to study the breakdown mechanisms and to develop new electrolytes for high operating voltage capacitors.


Laparoscopic Triple Segmental Bowel Resection for Endometriosis Revealed by Rectal Obstruction during Infertility Treatment.
Intestinal endometriosis is a benign disease characterized by ectopic growth of the endometrium and causes extensive fibrosis and adhesions in response to repeated episodes of bleeding and inflammation with the menstrual cycle. We encountered a rare case of intestinal endometriosis that caused complete rectal obstruction in a 34-year-old woman undergoing infertility treatment. Colonoscopy showed rectal stenosis and obstruction but no evidence of a tumor. Bowel obstruction due to endometriosis was diagnosed based on the history and imaging findings. Transanal decompression was performed. Subsequent laparoscopic surgery revealed severe inflammation around both ovaries and a tumor-like rectal stenosis. Similar findings were obtained in the transverse colon and terminal ileum. We performed laparoscopic low anterior resection, partial transverse colon resection, ileocecal resection, bilateral cystectomy, and left salpingectomy. Infertility treatment was restarted and resulted in a successful term pregnancy. The patient remains well. Laparoscopic surgery, which has the advantage of being minimally invasive, allows for early postoperative recovery and discharge in patients with endometriosis; furthermore, the uterus and adnexa can be preserved due to the magnifying effect of the laparoscope. In this case, it was possible to resume infertility treatment. Intestinal endometriosis is a rare cause of bowel obstruction, but should be kept in mind if intestinal obstruction occurs during infertility treatment. Laparoscopic surgery may be useful for multiple endometriotic lesions and serve as a bridge to infertility treatment.


Molecular cytogenetic (FISH) and genome analysis of diploid wheatgrasses and their phylogenetic relationship.
This paper reports detailed FISH-based karyotypes for three diploid wheatgrass species Agropyron cristatum (L.) Beauv., Thinopyrum bessarabicum (Savul.&Rayss) A. Löve, Pseudoroegneria spicata (Pursh) A. Löve, the supposed ancestors of hexaploid Thinopyrum intermedium (Host) Barkworth & D.R.Dewey, compiled using DNA repeats and comparative genome analysis based on COS markers. Fluorescence in situ hybridization (FISH) with repetitive DNA probes proved suitable for the identification of individual chromosomes in the diploid JJ, StSt and PP genomes. Of the seven microsatellite markers tested only the (GAA)n trinucleotide sequence was appropriate for use as a single chromosome marker for the P. spicata AS chromosome. Based on COS marker analysis, the phylogenetic relationship between diploid wheatgrasses and the hexaploid bread wheat genomes was established. These findings confirmed that the J and E genomes are in neighbouring clusters.


ECG changes in pneumothorax. A unique finding and proposed mechanism.
A patient with anterior myocardial infarction had bilateral pneumothoraces. Left pneumothorax was associated with reappearance of precordial R waves, while right pneumothorax caused no ECG change. This previously undescribed finding suggests the importance of air insulating the chest wall, rather than cardiac rotation, dilatation, or displacement as a mechanism of the ECG change.


An approach to defining and achieving restoration targets for a threatened plant community.
Connecting scientific research and government policy is essential for achieving objectives in sustaining biodiversity in an economic context. Our approach to connecting theoretical ecology, applied ecology, and policy was devised using principles of restoration ecology and the requisite methodology to restore biodiverse ecosystems. Using a threatened ecological community (TEC) with >120 plant species, we posit our approach as a guide for interpreting and achieving regulatory compliance (i.e., government conditions) enacted to manage or offset environmental impacts of development. We inform the scientific approach necessary to delivering outcomes appropriate to policy intent and biodiverse restoration through theoretical and applied research into the ecological restoration of the highly endemic flora of banded ironstone formations of the Mid West of Western Australia. Our approach (1) defines scale-appropriate restoration targets that meet regulatory compliance (e.g., Government of Western Australia Ministerial Conditions); (2) determines the optimal method to return individual plant species to the restoration landscape; (3) develops a conceptual model for our system, based on existing restoration frameworks, to optimize and facilitate the pathway to the restoration of a vegetation community (e.g., TEC) using diverse research approaches; and (4) develops an assessment protocol to compare restoration achievements against the expected regulatory outcomes using our experimental restoration trials as a test example. Our approach systematically addressed the complex challenges in setting and achieving restoration targets for an entire vegetation community, a first for a semiarid environment. We interpret our approach as an industry application relevant to policy- or regulator-mediated mine restoration programs that seek to return biodiverse species assemblages at landscape scales.


Heterologous expression of mating-type genes in filamentous fungi.
Podospora anserina and Neurospora crassa, two filamentous heterothallic ascomycetes, have a single mating-type locus with two alternate forms called mat+ and mat- and A and a, respectively. Mating type controls entry into the sexual cycle, events subsequent to fertilization, and, in N. crassa, prevents the formation of mixed mating-type heterokaryons. The mating types of these two organisms display similarity in their DNA structure and in the encoded polypeptides involved in fertilization. Here we show that this molecular similarity reflects a functional homology with respect to mating identity. Transformation experiments show that the N. crassa mating-type genes can provide the fertilization functions in P. anserina strains devoid of mating specificity as well as in mat+ and mat- strains. Reciprocally, the introduction of P. anserina mating-type genes confers mating activity in N. crassa. Functional identity between the mating types is not observed for vegetative incompatibility or for post-fertilization events such as meiosis and ascosporogenesis.


Electrical pulsed stimulation of surfaces homogeneously coated with gold nanoparticles to induce neurite outgrowth of PC12 cells.
In this study, the potential of gold nanoparticles (20 nm) to deliver electrical stimulation to nerve cell cultures in vitro to induce nerve regeneration was evaluated. In order to use these biomaterials to deliver an electrical stimulus, we devised a novel method for the fabrication of a nanostructured 2D substrate comprising gold nanoparticles attached to the surface of a cover glass via an adsorption system. In this strategy, gold nanoparticles are created and then coated onto a positively charged cover glass that has been pretreated with polyethyleneimine (PEI). Scanning electron microscopy (SEM) revealed that the PC 12 cells extended neurites well on the gold nanoparticles in the presence of electrical stimulation. In addition, the neurite outgrowth of PC12 cells in response to pulsed and constant electrical stimulation was evaluated by live/dead cell determination, by reverse transcriptase polymerase chain reaction (RT-PCR) analysis, as well as by beta-tubulin and NF-200 expression. By electrical stimulation (250 mV for 1 h), PC12 cells with their neurite outgrowth length were highly increased, with a mean diameter of 98.5 microm. However, the neurite outgrowth length without electrical stimulation was approximately 10 approximately 20 microm. Moreover, the alternating current stimulation also showed good viability (<90%), while a high amount of cell death (more than 30%) was observed with constant current stimulation. Thus, the gold nanoparticles with pulsed current stimulation may provide suitable tools for the nerve regeneration using neuronal cells.


On the absence of collective motion in a bulk suspension of spontaneously rotating dielectric particles.
A suspension of dielectric particles rotating spontaneously when subjected to a DC electric field in two dimensions next to a no-slip electrode has proven to be an ideal model for active matter [Bricard et al., Nature, 2013, 503, 95-98]. In this system, an electrohydrodynamic (EHD) instability called Quincke rotation was exploited to create self-propelling particles which aligned with each other due to EHD interactions, giving rise to collective motion on large length scales. It is natural to question whether a suspension of such particles in three dimensions will also display collective motion and spontaneously flow like bacterial suspensions do. Using molecular dynamics type simulations, we show that dielectrophoretic forces responsible for chaining in the direction of the applied electric field in conventional electrorheological fluids and the counter-rotation of neighboring particles in these chains prevent collective motion in suspensions undergoing spontaneous particle rotations. Our simulations discover that the fundamental microstructural unit of a suspension under Quincke rotation is a pair of counter-rotating spheres aligned in the direction of the electric field. We perform a linear stability analysis that explains this observation.


In-home consumer and shear force evaluation of steaks from the M. serratus ventralis thoracis.
The M. serratus ventralis thoracis was obtained from US Select arm chucks (n=87) to investigate if this underutilized muscle can be used as a steak alternative. Muscles were assigned randomly into three treatment groups: (1) control; (2) blade tenderization; and (3) injection, containing salt, phosphate, and papain. Steaks were cut from each muscle for in-home consumer evaluation (n=136) and Warner-Bratzler shear (WBS) force determination. The WBS values for injected steaks (13.1N) were lower (P<0.05) than for blade-tenderized (18.4N) and control (19.9N) steaks. Tenderness ratings for the injected steaks were higher (P<0.05) compared to the other treatments when steaks were grilled, oven prepared or were cooked in a skillet; however, this improvement did not significantly influence overall like scores for steaks that were oven prepared or cooked in a skillet. For the most part, degree of doneness did not significantly impact consumer evaluations of steaks prepared by the various cooking methods. However, there was a treatment x degree of doneness interaction for grilled-cooked steaks where increased doneness for blade-tenderized and injected steaks resulted in increased palatability ratings, whereas increased doneness for control steaks generally resulted in lowered palatability ratings. Consumer ratings and WBS values for the M. serratus ventralis thoracis indicate that merchandising steaks from this muscle may be a viable option in the marketplace, especially if blade tenderization or injection processes are used for further enhancement.


Gabapentin activates presynaptic GABAB heteroreceptors in rat cortical slices.
In electrically stimulated rat neocortical brain slices preloaded with [3H]gamma-aminobutyric acid (GABA) or [3H]glutamic acid, the pharmacological actions of 1-(aminomethyl)-cyclohexaneacetic acid (gabapentin, Gp) were compared with the GABAB receptor agonists baclofen (Bac) and (3-amino-2-(S)-hydroxypropyl)-methylphosphinic acid (CGP 44532). Gabapentin, baclofen and CGP 44532 all reduced the electrically stimulated release of [3H]glutamic acid (IC50=20 microM, 0.8 microM and 2 microM, respectively). These effects were sensitive to the GABAB receptor antagonists (+)-(S)-5,5 dimethylmorpholinyl-2-acetic acid (Sch 50911) or N-3-[[1-(S)-(3,4-dichlorophenyl)ethyl]amino]-2-(S)-hydroxypropyl-P-(cyclo-hexylmethyl)-phosphinic acid (CGP 54626). By contrast, gabapentin was without effect on the release of [3H]GABA, whilst baclofen (IC50=8 microM) and CGP 44532 (IC50=1 microM) inhibited [3H]GABA release. It is concluded that gabapentin selectively activates presynaptic GABAB heteroreceptors, but not GABAB autoreceptors, and may be a useful ligand to discriminate between presynaptic GABAB receptor subtypes.


<Editors' Choice> Initial experience with in vivo "endovascular shaping" technique that utilizes the vascular configuration for small cerebral aneurysm coiling.
Optimal shaping of a microcatheter tip is a key factor in cerebral aneurysm coiling. However, safe cannulation and stabilization of a microcatheter can be technically challenging with small aneurysms requiring precise microcatheter shaping. We devised a new microcatheter shaping technique which bends a microcatheter tip in the intended direction by placing and keeping the tip in the selected branch of the parent artery for 5 minutes: endovascular shaping technique. Our method can complete microcatheter shaping only inside the patient's body; an endovascularly shaped microcatheter will never face a straightening deformity and rotation problem associated with catheter reinsertion. The aim was to assess the feasibility and safety of endovascular shaping for small aneurysm coiling. Clinically, 10 consecutive challenging small terminal-type aneurysms (< 5 mm) treated with endovascular shaping were included. We retrospectively analyzed the newly acquired bend angle of a microcatheter tip after the shaping, the procedural success, and clinical outcomes. An artificial vascular model was used to confirm our findings. In all the 10 patients (three middle cerebral artery, four anterior communicating artery, and three basilar artery aneurysms), the endovascularly shaped microcatheters were bent toward the placement branch at an average of 21.7° and excellently guided into the aneurysms. Coil embolization was successfully accomplished without any complications. The results of the vascular model experiment demonstrated that an endovascularly shaped microcatheter acquired new bend angle toward the placement branch. Endovascular shaping was feasible and safe for small terminal-type aneurysm coiling. Our method can provide accurate shaping and stability during aneurysm coiling procedure.


[Sonographic imaging and staging of colonic tumors].
Out of 25 clinically, endoscopically or radiologically assured colonic carcinomas 84% could be imaged by sonography and 71% of those could be staged correctly. There were no differences in imaging of carcinomas between different parts of the colon. Sonographic detection of lymphnodules was only partially successful, whereas metastases of the carcinomas were shown in 6 out of 7 cases. Thus sonographic investigation of the colon may provide non-invasive staging of an assured colonic carcinoma and its follow-up controls.


The D2R-DISC1 protein complex and associated proteins are altered in schizophrenia and normalized with antipsychotic treatment.
For decades, the dopamine D2 receptor (D2R) has been known as the main target of antipsychotic medications, but the mechanism for antipsychotic effects beyond this pharmacological target remains unclear. Disrupted-in-schizophrenia 1 (DISC1) is a gene implicated in the etiology of schizophrenia, and we have found elevated levels of the D2R-DISC1 complex in the postmortem brain tissue of patients with schizophrenia. We used coimmunoprecipitation to measure D2R-DISC1 complex levels in peripheral blood samples from patients with schizophrenia and unaffected controls in 3 cohorts (including males and females) from different hospitals. We also used label-free mass spectrometry to conduct proteomic analysis of these samples. Levels of the D2R-DISC1 complex were elevated in the peripheral blood samples of patients with schizophrenia from 3 independent cohorts, and were normalized with antipsychotic treatment. Proteomic analysis of the blood samples from patients with high D2R-DISC1 complex levels that were normalized with antipsychotic treatment revealed a number of altered proteins and pathways associated with D2R, DISC1 and the D2R-DISC1 complex. We identified additional proteins and pathways that were associated with antipsychotic treatment in schizophrenia, and that may also be novel targets for schizophrenia treatment. Sample sizes were relatively small, but were sufficient to detect associations between D2R-DISC1 levels, schizophrenia and treatment response. The relevance of leukocyte changes to the symptoms of schizophrenia is unknown. The coimmunoprecipitation lanes included several nonspecific bands. Levels of the D2R-DISC1 complex were elevated in patients with schizophrenia and reduced with antipsychotic treatment. This finding reinforces the independent role of each protein in schizophrenia. Our results enhanced our understanding of the molecular pathways involved in schizophrenia and in antipsychotic medications, and identified novel potential molecular targets for treating schizophrenia.


Aberrant methylation and downregulation of sall3 in human hepatocellular carcinoma.
To investigated whether sall3 transcription was regulated by promoter CpG island hypermethylation in hepatocellular carcinoma (HCC). The cell lines Huh7, HepG2, SK-HEP1, SMMC7721, Bel7402, QGY7703 and a cohort of 38 HCC tissue specimens and corresponding nontumorous tissues were subjected to analysis for sall3 promoter CpG island methylation and mRNA transcription. sall3 promoter CpG island methylation levels were determined using the MassARRAY platform and mRNA transcription levels of the gene were detected by quantitative real-time polymerase chain reaction. The levels of sall3 mRNA were decreased by more than twofold in 33 of 38 tumor tissues compared to adjacent noncancerous tissues. Among these 33 tumor tissues with lower levels of sall3 mRNA, 24 showed higher levels of methylation. Based on these results, we hypothesized that the decrease in sall3 mRNA transcription level was likely due to promoter CpG island hypermethylation. Changes in sall3 mRNA transcription and promoter CpG island methylation were determined in the above six cell lines after treatment with 0, 0.1, 0.5 and 2.5 μmol 5-aza-2-deoxycytidine, a demethylating agent. Promoter CpG island methylation levels decreased in a dose-dependent manner in all six cell lines, while the mRNA transcription level increased dose-dependently in Huh7, HepG2, SK-HEP1 and SMMC7721 cells and irregularly in Bel7402 and QGY7703 cells. These results indicated that promoter CpG island hypermethylation contributes to the downregulation of sall3 mRNA transcription in HCC.


Analysis of recruitment in a pragmatic observational study on C-reactive protein point-of-care testing in primary care.
Failure to recruit all eligible study patients can lead to biased results. Little is known on selective patient recruitment in studies on implementation of diagnostic devices. The aim of this observational study was to measure recruitment of patients in an implementation study in primary care on use of point-of-care (POC) C-reactive protein (CRP) and to evaluate recruitment bias and its impact on the study endpoint. In a cross-sectional observational study on POC CRP implementation and related antibiotics prescribing, we compared included patients with all eligible patients to assess the representativeness of the included subjects. Eligible patients were adults presenting with acute cough in primary care between March and September 2012. The frequency of POC CRP testing and the proportion of prescribed antibiotics were compared between recruited and non-recruited patients. As measure of bias, odds ratios (ORs) with accompanying 95% confidence intervals (CIs) for the association between CRP level (<20 mg/l or not) and antibiotic prescribing were computed. Of all 1473 eligible patients 348 (24%) were recruited. In recruited patients, POC CRP tests were conducted and antibiotics prescribed more frequently as compared to non-recruited patients (81% versus 6% and 44% versus 29%, respectively). The ORs were 18.2 (95%CI: 9.6-34.3), 30.5 (95%CI: 13.2-70.3) and 3.8 (95%CI: 0.9-14.8) respectively in all eligible patients, the recruited and the non-recruited patients. Selective recruitment resulted in an overestimation of POC CRP test use and antibiotic prescribing.