New approach to capture and characterize synaptic proteome

Little is known regarding the identity of the population of proteins that are transported and localized to synapses. Here we describe a new approach that involves the isolation and systematic proteomic characterization of molecular motor kinesins to identify the populations of proteins transported to synapses. We used this approach to identify and compare proteins transported to synapses by kinesin (Kif) complexes Kif5C and Kif3A in the mouse hippocampus and prefrontal cortex. Approximately 40–50% of the protein cargos identified in our proteomics analysis of kinesin complexes are known synaptic proteins. We also found that the identity of kinesins and where they are expressed determine what proteins they transport. Our results reveal a previously unappreciated role of kinesins in regulating the composition of synaptic proteome.A number of researchers have reported specific roles of proteins that are localized to synapses, termed the “synaptic proteome,” in determining synaptic function and plasticity (1–6). Identifying the composition and dynamics of the synaptic proteome is key to understanding the remodeling of existing synapses and growth of new synapses, as well as identifying novel therapeutic targets for synaptopathies (7, 8). The composition of the synaptic proteome is determined by proteins transported from the cell body and local synthesis of proteins at the synapses. Several studies have shown the identification and localization of RNAs at the synapse (9–12). Comprehensive knowledge of the proteins that are actively targeted to synapses will help elucidate the dynamics of signal transduction at the synapse, as well as the molecules that are critical for the remodeling of synapses and formation of new synapses that accompany long-term memory storage. Despite this perceived significance, however, there are no currently available methodologies for studying populations of proteins targeted to synapses.To identify actively transported proteins, we focused on the protein complexes associated with kinesins, molecular motor proteins that move from the cell body to distal neuronal processes and mediate the transport of various gene products (9, 13). We considered that proteins associated with kinesins include both synaptic proteins and proteins localized to dendrites and axons. Our methodology involved isolating specific kinesin complexes by coimmunoprecipitation (co-IP) from different regions of the mouse brain and systematically identifying the protein cargos by mass spectrometry (Fig. 1A). Using this strategy, we have successfully isolated and characterized the Kif5C kinesin complex from hippocampus and prefrontal cortex (PFC) and the Kif3A kinesin complex from hippocampus. The cargos include organelles and proteins involved in translation, signaling, and ion channels. Several of these proteins are implicated in neuropsychiatric disorders (14).Open in a separate windowFig. 1.Identification of the Kif5C complex in mouse hippocampus. (A) Strategy used to analyze Kif5C-specific protein cargos in the mouse hippocampus. The schematic diagram shows the workflow for profiling Kif5C-transported proteins by proteomics. The processes include brain dissection, hippocampal tissue extraction and homogenization, Kif5C IP, complex separation by SDS/PAGE, isolation of specific bands followed by in-gel tryptic digestion, and LC-MS/MS analysis. (B) IP of Kif5C from mouse hippocampus. Kif5C complexes were immunoprecipitated from hippocampus using anti Kif5C antibody. (Upper) Protein complexes were then separated on SDS/PAGE and stained with silver. (Lower) Kif5C also was detected by Western blot analysis. Immunoprecipitation using CREB antibody and beads alone served as controls for IP. (C) Co-IP and Western blot analysis of Kif5C cargo proteins (PURα, DIC1, and GluR1) in mouse hippocampus. Total extract (input), IP, post-IP supernatant, beads-alone IP, and post-IP supernatant are shown. Arrows in B indicate the positions of Kif5C and antibody.     

Exploring the potential of a conditional cash transfer intervention to reduce HIV risk among young women in Iringa,Tanzania

Cash transfer programs seek to alter structural determinants of HIV risk such as poverty and gender inequality. We sought to explore the feasibility and potential effectiveness of a cash transfer intervention for young women as part of combination HIV prevention in Iringa, Tanzania. Qualitative, in-depth interviews were conducted with 116 stakeholders and residents from the region, including key informants, service delivery users, and members of key populations. Most respondents felt a cash transfer program would assist young women in Iringa to have more control over sexual decision-making and reduce poverty-driven transactional sex. Respondents were divided on who should receive funds: young women themselves, their parents/guardians, or community leaders. Cash amounts and suggested target groups varied, and several respondents suggested providing microcredit or small business capital instead of cash. Potential concerns included jealousy, dependency, and corruption. However, most respondents felt that some intervention was needed to address underlying poverty driving some sexual risk behavior. A cash transfer program could fill this role, ultimately reducing HIV, sexually transmitted infections, and unintended pregnancies. As increased attention is given to economic and structural interventions for HIV prevention, local input and knowledge should be considered in a program design.     

Differences in the right inferior longitudinal fasciculus but no general disruption of white matter tracts in children with autism spectrum disorder

One of the most widely cited features of the neural phenotype of autism is reduced “integrity” of long-range white matter tracts, a claim based primarily on diffusion imaging studies. However, many prior studies have small sample sizes and/or fail to address differences in data quality between those with autism spectrum disorder (ASD) and typical participants, and there is little consensus on which tracts are affected. To overcome these problems, we scanned a large sample of children with autism (n = 52) and typically developing children (n = 73). Data quality was variable, and worse in the ASD group, with some scans unusable because of head motion artifacts. When we follow standard data analysis practices (i.e., without matching head motion between groups), we replicate the finding of lower fractional anisotropy (FA) in multiple white matter tracts. However, when we carefully match data quality between groups, all these effects disappear except in one tract, the right inferior longitudinal fasciculus (ILF). Additional analyses showed the expected developmental increases in the FA of fiber tracts within ASD and typical groups individually, demonstrating that we had sufficient statistical power to detect known group differences. Our data challenge the widely claimed general disruption of white matter tracts in autism, instead implicating only one tract, the right ILF, in the ASD phenotype.What is the key difference in the brains of individuals with autism that accounts for the distinctive cognitive profile of this disorder? One of the most widely claimed brain signatures of autism spectrum disorder (ASD), reported in dozens of papers that used diffusion-weighted imaging (DWI), is reduced integrity of long-range fiber tracts (1). This finding has been taken as evidence that autism is fundamentally a “disconnection” syndrome, in which the core cognitive deficits result from reduced integration of information at the neural and cognitive levels (2–5). For example, it has been argued that the characteristic deficits in social cognition and language arise because these functions require rapid integration of information across spatially distant brain areas (3, 6, 7), which would likely be affected if major white matter tracts are compromised.Evidence for a general reduction in the “integrity”^* of white matter in autism has come primarily from diffusion imaging studies that report reduced directionality of the diffusion of water molecules, or fractional anisotropy (FA), and increased speed of diffusion, or mean diffusivity (MD) of many major fiber bundles. However, the literature reveals little actual agreement on the existence and direction of group differences in diffusion parameters (reviewed in ref. 1). White-matter differences have been reported in various brain regions in positive and negative directions. Possible reasons for these inconsistent findings include small sample sizes [mean of ∼20 in each group, with 40% of studies scanning 15 or fewer participants with ASD (1)], the heterogeneity of ASD itself, variations across studies in the age of the cohort tested, and the type of DTI analysis performed. Another potential problem that few diffusion studies of autism address or even mention is data quality. Indeed, to our knowledge, only two studies (9, 10) report quantitative analyses of the amount of motion in their DWI data. Group differences in head motion could be a serious confounding factor, given that head motion is likely to be greater in children with autism, and group differences in head motion can lead artifactually to just the effects most often reported: reduced FA in white matter tracts in ASD (11).To address these concerns, we scanned a relatively large sample of children with and without ASD, and evaluated data quality from each participant by visual inspection of the data and quantification of head motion (11). We then excluded scans that did not reach our data quality criterion, and matched the remaining participants across groups for data quality. These data were used to determine whether people with autism do in fact show widespread differences in the known white matter tracts in ASD. We further tested the specific hypothesis that individuals with ASD show changes in one particular tract, the inferior longitudinal fasciculus (ILF), a white matter tract important for face recognition (12, 13), a mental function selectively disrupted in ASD (ref. 14; but see ref. 15).     

Vital Signs: Fruit and Vegetable Intake Among Children — United States, 2003–2010

Background

Eating more fruits and vegetables adds underconsumed nutrients to diets, reduces the risks for leading causes of illness and death, and helps manage body weight. This report describes trends in the contributions of fruits and vegetables to the diets of children aged 2–18 years.

Methods

CDC analyzed 1 day of 24-hour dietary recalls from the National Health and Nutrition Examination Surveys from 2003 to 2010 to estimate trends in children’s fruit and vegetable intake in cup-equivalents per 1,000 calories (CEPC) and trends by sex, age, race/ethnicity, family income to poverty ratio, and obesity status. Total fruit includes whole fruit (all fruit excluding juice) and fruit juice (from 100% juice, foods, and other beverages). Total vegetables include those encouraged in the Dietary Guidelines for Americans, 2010 (i.e., dark green, orange, and red vegetables and legumes), white potatoes, and all other vegetables.

Results

Total fruit intake among children increased from 0.55 CEPC in 2003–2004 to 0.62 in 2009–2010 because of significant increases in whole fruit intake (0.24 to 0.40 CEPC). Over this period, fruit juice intake significantly decreased (0.31 to 0.22 CEPC). Total vegetable intake did not change (0.54 to 0.53 CEPC). No socio-demographic group met the Healthy People 2020 target of 1.1 CEPC vegetables, and only children aged 2–5 years met the target of 0.9 CEPC fruits.

Conclusions

Children’s total fruit intake increased because of increases in whole fruit consumption, but total vegetable intake remained unchanged.

Implications for Public Health Practice

Increased attention to the policies and food environments in multiple settings, including schools, early care and education, and homes might help continue the progress in fruit intake and improve vegetable intake.     

A Randomized,Controlled Pragmatic Trial of Telephonic Medication Therapy Management to Reduce Hospitalization in Home Health Patients

Objective

To evaluate the effectiveness of a telephonic medication therapy management (MTM) service on reducing hospitalizations among home health patients.

Setting

Forty randomly selected, geographically diverse home health care centers in the United States.

Design

Two-stage, randomized, controlled trial with 60-day follow-up. All Medicare- insured home health care patients were eligible to participate. Twenty-eight consecutive patients within each care center were recruited and randomized to usual care or MTM intervention. The MTM intervention consisted of the following: (1) initial phone call by a pharmacy technician to verify active medications; (2) pharmacist-provided medication regimen review by telephone; and (3) follow-up pharmacist phone calls at day seven and as needed for 30 days. The primary outcome was 60-day all-cause hospitalization.

Data Collection

Data were collected from in-home nursing assessments using the OASIS-C. Multivariate logistic regression modeled the effect of the MTM intervention on the probability of hospitalization while adjusting for patients’ baseline risk of hospitalization, number of medications taken daily, and other OASIS-C data elements.

Principal Findings

A total of 895 patients (intervention n = 415, control n = 480) were block-randomized to the intervention or usual care. There was no significant difference in the 60-day probability of hospitalization between the MTM intervention and control groups (Adjusted OR: 1.26, 95 percent CI: 0.89–1.77, p = .19). For patients within the lowest baseline risk quartile (n = 232), the intervention group was three times more likely to remain out of the hospital at 60 days (Adjusted OR: 3.79, 95 percent CI: 1.35–10.57, p = .01) compared to the usual care group.

Conclusions

This MTM intervention may not be effective for all home health patients; however, for those patients with the lowest-risk profile, the MTM intervention prevented patients from being hospitalized at 60 days.     

Rapid dopamine transmission within the nucleus accumbens: Dramatic difference between morphine and oxycodone delivery

While most drugs of abuse increase dopamine neurotransmission, rapid neurochemical measurements show that different drugs evoke distinct dopamine release patterns within the nucleus accumbens. Rapid changes in dopamine concentration following psychostimulant administration have been well studied; however, such changes have never been examined following opioid delivery. Here, we provide novel measures of rapid dopamine release following intravenous infusion of two opioids, morphine and oxycodone, in drug‐naïve rats using fast‐scan cyclic voltammetry and rapid (1 min) microdialysis coupled with high‐performance liquid chromatography ‐ tandem mass spectrometry (HPLC‐MS). In addition to measuring rapid dopamine transmission, microdialysis HPLC‐MS measures changes in GABA, glutamate, monoamines, monoamine metabolites and several other neurotransmitters. Although both opioids increased dopamine release in the nucleus accumbens, their patterns of drug‐evoked dopamine transmission differed dramatically. Oxycodone evoked a robust and stable increase in dopamine concentration and a robust increase in the frequency and amplitude of phasic dopamine release events. Conversely, morphine evoked a brief (~ 1 min) increase in dopamine that was coincident with a surge in GABA concentration and then both transmitters returned to baseline levels. Thus, by providing rapid measures of neurotransmission, this study reveals previously unknown differences in opioid‐induced neurotransmitter signaling. Investigating these differences may be essential for understanding how these two drugs of abuse could differentially usurp motivational circuitry and powerfully influence behavior.     

Regulatory T-cells in helminth infection: induction,function and therapeutic potential

Helminth parasites infect an alarmingly large proportion of the world's population, primarily within tropical regions, and their ability to down-modulate host immunity is key to their persistence. Helminths have developed multiple mechanisms that induce a state of hyporesponsiveness or immune suppression within the host; of particular interest are mechanisms that drive the induction of regulatory T-cells (Tregs). Helminths actively induce Tregs either directly by secreting factors, such as the TGF-β mimic Hp-TGM, or indirectly by interacting with bystander cell types such as dendritic cells and macrophages that then induce Tregs. Expansion of Tregs not only enhances parasite survival but, in cases such as filarial infection, Tregs also play a role in preventing parasite-associated pathologies. Furthermore, Tregs generated during helminth infection have been associated with suppression of bystander immunopathologies in a range of inflammatory conditions such as allergy and autoimmune disease. In this review, we discuss evidence from natural and experimental infections that point to the pathways and molecules involved in helminth Treg induction, and postulate how parasite-derived molecules and/or Tregs might be applied as anti-inflammatory therapies in the future.     

Cytogenetic analysis of 130 renal oncocytomas identify three distinct and mutually exclusive diagnostic classes of chromosome aberrations

The cytogenetic alterations in renal oncocytoma (RO) are poorly understood. We analyzed 130 consecutive RO for karyotypic alterations. Clonal chromosome abnormalities were identified in 63 (49%) cases, which could be categorized into three classes of mutually exclusive cytogenetic categories. Class 1 (N = 20) RO had diploid karyotypes with characteristic 11q13 rearrangement in balanced translocations with 10 or more different chromosome partners in all cases. We identified recurrent translocation partners at 5q35, 6p21, 9p24, 11p13‐14, and 11q23, and confirmed that CCND1 gene rearrangement at 11q13 utilizing fluorescence in situ hybridization (FISH). Class 2 RO (N = 25) exhibited hypodiploid karyotypes with loss of chromosome 1 and/or losses of Y in males and X in females in all cases. The class 3 tumors comprising of 18 cases showed diverse types of abnormalities with the involvement of two or more chromosomes exclusive of abnormalities seen in classes 1 and 2 tumors. Furthermore, karyotypically uninformative cases were subjected to FISH analysis to identify classes 1 and 2 abnormalities. In this group, we found similar frequencies of CCND1 rearrangement, loss of chromosome 1 or Y as with karyotypically abnormal cases. We validated our results against 91 tumors from the Mitelman database. Correlation of clinical data with all the three classes of ROs showed no clear evidence of overall patient survival. Our findings support the hypothesis that RO exhibit three principal cytogenetic categories, which may have different roles in initiation and/or progression. These cytogenetic markers provide a key tool in the diagnostic evaluation of RO.     

Variation in accessory mental foramen frequency and number in extant hominoids

Nerves providing sensation to the lower face and jaw exit the mandibular canal via the mental foramen. In humans, there are many documented occurrences of additional foramina (accessory mental foramina, AMFs) on the lateral mandibular surface that may also contain nervous structures. There are large discrepancies in the literature regarding how often AMFs occur in humans, and investigations of non-human hominoid AMFs are rare. Consequently, the causes of interspecific diversity in this variable have not been explored. This project seeks to compare the frequency and number of AMFs between males and females, and among human regional groups and hominoid subspecies and species, and to investigate possible causal factors for any differences identified. No significant differences were found between males and females in any group. Gorillas and orangutans had the highest percentages of individuals with AMFs and the highest mean number of foramina, while modern humans and siamangs had the lowest figures for these variables. Significant differences (p < .05) were found for the mean number of foramina between most pairs of species. The results also showed that species with mandibles that are larger overall, have a larger area anterior to mental foramen, and a longer mandibular canal typically present more AMFs. The strongest correlation was found between the mean number of mental foramina and mandibular canal length. We suggest that these results provide preliminary support for the hypothesis that increasing mandibular canal length increases the likelihood that that nerves will ramify, leading to greater frequencies of accessory mental foramina.     

PEER umbrella systematic review of systematic reviews: Management of osteoarthritis in primary care

ObjectiveTo determine how many patients with chronic osteoarthritis pain respond to various non-surgical treatments.Data sourcesPubMed and the Cochrane Library.Study selection Published systematic reviews of randomized controlled trials (RCTs) that included meta-analysis of responder outcomes for at least 1 of the following interventions were included: acetaminophen, oral nonsteroidal anti-inflammatory drugs (NSAIDs), topical NSAIDs, serotonin-norepinephrine reuptake inhibitors (SNRIs), tricyclic antidepressants, cannabinoids, counseling, exercise, platelet-rich plasma, viscosupplementation, glucosamine, chondroitin, intra-articular corticosteroids, rubefacients, or opioids.Synthesis In total, 235 systematic reviews were included. Owing to limited reporting of responder meta-analyses, a post hoc decision was made to evaluate individual RCTs with responder analysis within the included systematic reviews. New meta-analyses were performed where possible. A total of 155 RCTs were included. Interventions that led to more patients attaining meaningful pain relief compared with control included exercise (risk ratio [RR] of 2.36; 95% CI 1.79 to 3.12), intra-articular corticosteroids (RR = 1.74; 95% CI 1.15 to 2.62), SNRIs (RR = 1.53; 95% CI 1.25 to 1.87), oral NSAIDs (RR = 1.44; 95% CI 1.36 to 1.52), glucosamine (RR = 1.33; 95% CI 1.02 to 1.74), topical NSAIDs (RR = 1.27; 95% CI 1.16 to 1.38), chondroitin (RR = 1.26; 95% CI 1.13 to 1.41), viscosupplementation (RR = 1.22; 95% CI 1.12 to 1.33), and opioids (RR = 1.16; 95% CI 1.02 to 1.32). Preplanned subgroup analysis demonstrated no effect with glucosamine, chondroitin, or viscosupplementation in studies that were only publicly funded. When trials longer than 4 weeks were analyzed, the benefits of opioids were not statistically significant.ConclusionInterventions that provide meaningful relief for chronic osteoarthritis pain might include exercise, intra-articular corticosteroids, SNRIs, oral and topical NSAIDs, glucosamine, chondroitin, viscosupplementation, and opioids. However, funding of studies and length of treatment are important considerations in interpreting these data.