‘Fractional Recovery’ Analysis of a Presynaptic Synaptotagmin 1-Anchored Endocytic Protein Complex

BACKGROUND: The integral synaptic vesicle protein and putative calcium sensor, synaptotagmin 1 (STG), has also been implicated in synaptic vesicle (SV) recovery. However, proteins with which STG interacts during SV endocytosis remain poorly understood. We have isolated an STG-associated endocytic complex (SAE) from presynaptic nerve terminals and have used a novel fractional recovery (FR) assay based on electrostatic dissociation to identify SAE components and map the complex structure. The location of SAE in the presynaptic terminal was determined by high-resolution quantitative immunocytochemistry at the chick ciliary ganglion giant calyx-type synapse. METHODOLOGY/PRINCIPLE FINDINGS: The first step in FR analysis was to immunoprecipitate (IP) the complex with an antibody against one protein component (the IP-protein). The immobilized complex was then exposed to a high salt (1150 mM) stress-test that caused shedding of co-immunoprecipitated proteins (co-IP-proteins). A Fractional Recovery ratio (FR: recovery after high salt/recovery with control salt as assayed by Western blot) was calculated for each co-IP-protein. These FR values reflect complex structure since an easily dissociated protein, with a low FR value, cannot be intermediary between the IP-protein and a salt-resistant protein. The structure of the complex was mapped and a blueprint generated with a pair of FR analyses generated using two different IP-proteins. The blueprint of SAE contains an AP180/X/STG/stonin 2/intersectin/epsin core (X is unknown and epsin is hypothesized), and an AP2 adaptor, H-/L-clathrin coat and dynamin scission protein perimeter. Quantitative immunocytochemistry (ICA/ICQ method) at an isolated calyx-type presynaptic terminal indicates that this complex is associated with STG at the presynaptic transmitter release face but not with STG on intracellular synaptic vesicles. CONCLUSIONS/SIGNIFICANCE: We hypothesize that the SAE serves as a recognition site and also as a seed complex for clathrin-mediated synaptic vesicle recovery. The combination of FR analysis with quantitative immunocytochemistry provides a novel and effective strategy for the identification and characterization of biologically-relevant multi-molecular complexes

Leukotriene antagonists as first-line or add-on asthma controller therapy

Most randomized trials of treatment for asthma study highly selected patients under idealized conditions. METHODS: We conducted two parallel, multicenter, pragmatic trials to evaluate the real-world effectiveness of a leukotriene-receptor antagonist (LTRA) as compared with either an inhaled glucocorticoid for first-line asthma-controller therapy or a long-acting beta(2)-agonist (LABA) as add-on therapy in patients already receiving inhaled glucocorticoid therapy. Eligible primary care patients 12 to 80 years of age had impaired asthma-related quality of life (Mini Asthma Quality of Life Questionnaire [MiniAQLQ] score =6) or inadequate asthma control (Asthma Control Questionnaire [ACQ] score =1). We randomly assigned patients to 2 years of open-label therapy, under the care of their usual physician, with LTRA (148 patients) or an inhaled glucocorticoid (158 patients) in the first-line controller therapy trial and LTRA (170 patients) or LABA (182 patients) added to an inhaled glucocorticoid in the add-on therapy trial. RESULTS: Mean MiniAQLQ scores increased by 0.8 to 1.0 point over a period of 2 years in both trials. At 2 months, differences in the MiniAQLQ scores between the two treatment groups met our definition of equivalence (95% confidence interval [CI] for an adjusted mean difference, -0.3 to 0.3). At 2 years, mean MiniAQLQ scores approached equivalence, with an adjusted mean difference between treatment groups of -0.11 (95% CI, -0.35 to 0.13) in the first-line controller therapy trial and of -0.11 (95% CI, -0.32 to 0.11) in the add-on therapy trial. Exacerbation rates and ACQ scores did not differ significantly between the two groups. CONCLUSIONS: Study results at 2 months suggest that LTRA was equivalent to an inhaled glucocorticoid as first-line controller therapy and to LABA as add-on therapy for diverse primary care patients. Equivalence was not proved at 2 years. The interpretation of results of pragmatic research may be limited by the crossover between treatment groups and lack of a placebo group

Enhancement of nematic order and global phase diagram of a lattice model for coupled nematic systems

We use an infinite-range Maier-Saupe model, with two sets of local quadrupolar variables and restricted orientations, to investigate the global phase diagram of a coupled system of two nematic subsystems. The free energy and the equations of state are exactly calculated by standard techniques of statistical mechanics. The nematic-isotropic transition temperature of system A increases with both the interaction energy among mesogens of system B, and the two-subsystem coupling $J$. This enhancement of the nematic phase is manifested in a global phase diagram in terms of the interaction parameters and the temperature $T$. We make some comments on the connections of these results with experimental findings for a system of diluted ferroelectric nanoparticles embedded in a nematic liquid-crystalline environment.Comment: 11 pages, 3 figures, to appear in Volume 42 of the Brazilian Journal of Physic

Order cones: A tool for deriving k-dimensional faces of cones of subfamilies of monotone games

In this paper we introduce the concept of order cone. This concept is inspired by the concept of order polytopes, a well-known object coming from Combinatorics. Similarly to order polytopes, order cones are a special type of polyhedral cones whose geometrical structure depends on the properties of a partially ordered set (brief poset). This allows to study these properties in terms of the subjacent poset, a problem that is usually simpler to solve. From the point of view of applicability, it can be seen that many cones appearing in the literature of monotone TU-games are order cones. Especially, it can be seen that the cones of monotone games with restricted cooperation are order cones, no matter the structure of the set of feasible coalitions

Human papillomavirus and vaccination: knowledge, attitudes, and behavioural intention in adolescents and young women in Italy

This study assesses knowledge, attitudes, and behavioural intention towards human papillomavirus (HPV) infection and vaccination in a random sample of 1348 adolescents and young women aged 14–24 years in Italy. A self-administered anonymous questionnaire covered demographics; knowledge about HPV infection, cervical cancer, and HPV vaccine; the perceived risk for contracting HPV infection and/or for developing cervical cancer, the perceived benefits of a vaccination to prevent cervical cancer, and willingness to receive an HPV vaccine. Only 23.3% have heard that HPV is an infection of the genital mucosa and about cervical cancer. Those older, with at least one parent who is a health care professional, with personal, familiar, or friendly history of cervical cancer, and having underwent a health checkup in the last year with information about HPV vaccination were significantly more knowledgeable. Risk perception scores (range: 1–10) of contracting HPV infection and of developing cervical cancer were 5.8 and 6.5. Older age, not having a parent who is a health care professional, having had a personal, familiar, or friendly history of cervical cancer, and need of additional information were predictors of the perceived susceptibility of developing cervical cancer. The vast majority professed intent to receive an HPV vaccine and the significant predictors were having at least one parent who is a health care professional, a high perceived risk of contracting HPV infection and of developing cervical cancer, and a high belief towards the utility of a vaccination for preventing cervical cancer. Knowledge about HPV infection and cervical cancer should be improved with more attention to the benefit of HPV vaccination

Epidemiologic and clinical updates on impulse control disorders: a critical review

The article reviews the current knowledge about the impulse control disorders (ICDs) with specific emphasis on epidemiological and pharmacological advances. In addition to the traditional ICDs present in the DSM-IV—pathological gambling, trichotillomania, kleptomania, pyromania and intermittent explosive disorder—a brief description of the new proposed ICDs—compulsive–impulsive (C–I) Internet usage disorder, C–I sexual behaviors, C–I skin picking and C–I shopping—is provided. Specifically, the article summarizes the phenomenology, epidemiology and comorbidity of the ICDs. Particular attention is paid to the relationship between ICDs and obsessive–compulsive disorder (OCD). Finally, current pharmacological options for treating ICDs are presented and discussed

Body mass index is associated with reduced exhaled nitric oxide and higher exhaled 8-isoprostanes in asthmatics

BACKGROUND: Recently, it has been shown that increasing body mass index (BMI) in asthma is associated with reduced exhaled NO. Our objective in this study was to determine if the BMI-related changes in exhaled NO differ across asthmatics and controls, and to determine if these changes are related to increased airway oxidative stress and systemic levels of leptin and adiponectin. METHODS: Observational study of the association of BMI, leptin, and adiponectin with exhaled nitric oxide (NO) and exhaled 8-isoprostanes in 67 non-smoking patients with moderate to severe persistent asthma during baseline conditions and 47 controls. Measurements included plasma levels of leptin, adiponectin, exhaled breath condensates for 8-isoprostanes, exhaled NO, pulmonary function tests, and questionnaires regarding asthma severity and control. RESULTS: In asthmatics, BMI and the ratio of leptin to adiponectin were respectively associated with reduced levels of exhaled NO (β = -0.04 [95% C.I. -0.07, -0.1], p < 0.003) and (β = -0.0018 [95% C.I. -0.003, -0.00034], p = 0.01) after adjusting for confounders. Also, BMI was associated with increased levels of exhaled 8-isoprostanes (β = 0.30 [95% C.I. 0.003, 0.6], p = 0.03) after adjusting for confounders. In contrast, we did not observe these associations in the control group of healthy non-asthmatics with a similar weight distribution. CONCLUSION: In adults with stable moderate to severe persistent asthma, but not in controls, BMI and the plasma ratio of leptin/adiponectin is associated with reduced exhaled NO. Also, BMI is associated with increased exhaled 8-isoprostanes. These results suggest that BMI in asthmatics may increase airway oxidative stress and could explain the BMI-related reductions in exhaled NO

An Inhibitory Effect of Extracellular Ca2+ on Ca2+-Dependent Exocytosis

Aim: Neurotransmitter release is elicited by an elevation of intracellular Ca 2+ concentration ([Ca 2+] i). The action potential triggers Ca 2+ influx through Ca 2+ channels which causes local changes of [Ca 2+] i for vesicle release. However, any direct role of extracellular Ca 2+ (besides Ca 2+ influx) on Ca 2+-dependent exocytosis remains elusive. Here we set out to investigate this possibility on rat dorsal root ganglion (DRG) neurons and chromaffin cells, widely used models for studying vesicle exocytosis. Results: Using photolysis of caged Ca 2+ and caffeine-induced release of stored Ca 2+, we found that extracellular Ca 2+ inhibited exocytosis following moderate [Ca 2+]i rises (2–3 mM). The IC50 for extracellular Ca 2+ inhibition of exocytosis (ECIE) was 1.38 mM and a physiological reduction (,30%) of extracellular Ca 2+ concentration ([Ca 2+]o) significantly increased the evoked exocytosis. At the single vesicle level, quantal size and release frequency were also altered by physiological [Ca 2+] o. The calcimimetics Mg 2+,Cd 2+, G418, and neomycin all inhibited exocytosis. The extracellular Ca 2+-sensing receptor (CaSR) was not involved because specific drugs and knockdown of CaSR in DRG neurons did not affect ECIE. Conclusion/Significance: As an extension of the classic Ca 2+ hypothesis of synaptic release, physiological levels of extracellular Ca 2+ play dual roles in evoked exocytosis by providing a source of Ca 2+ influx, and by directly regulatin