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Children's mental health is deteriorating while access to child and adolescent mental health services is decreasing. Recent UK policy has focused on schools as a setting for the provision of mental health services, and counselling is the most common type of school-based mental health provision. This study examined the longer-term effectiveness of one-to-one school-based counselling delivered to children in UK primary schools. Data were drawn from a sample of children who received school-based counselling in the UK in the 2015/16 academic year, delivered by a national charitable organisation. Mental health was assessed at baseline, immediately post-intervention, and approximately 1 year post-intervention, using the Strengths and Difficulties Questionnaire (SDQ) completed by teachers and parents. Paired t tests compared post-intervention and follow-up SDQ total difficulties scores with baseline values. Propensity score matching was then used to identify a comparator group of children from a national population survey, and linear mixed effects models compared trajectories of SDQ scores in the two groups. In the intervention group, teacher and parent SDQ total difficulties scores were lower at post-intervention and longer-term follow-up compared to baseline (teacher: baseline 14.42 (SD 7.18); post-intervention 11.09 (6.93), t(739) = 13.78, p < 0.001; follow-up 11.27 (7.27), t(739) = 11.92, p < 0.001; parent: baseline 15.64 (6.49); post-intervention 11.90 (6.78), t(361 = 11.29, p < 0.001); follow-up 11.32 (7.19), t(361) = 11.29, p < 0.001). The reduction in SDQ scores was greater in the intervention compared to the comparator group (likelihood ratio test comparing models with time only versus time plus group-by-time interaction: χ2 (3) = 24.09, p < 0.001), and model-predicted SDQ scores were lower in the intervention than comparator group for 2 years post-baseline. A one-to-one counselling intervention delivered to children in UK primary schools predicted improvements in mental health that were maintained over a 2 year follow-up period.

Exclusion from school is a disciplinary tool with an increasingly recognised relationship to poor mental health among children and young people. We explored the relationship between mental health and school exclusion for a cohort of children and young people receiving one to one counselling.We analysed routinely collected data from a diverse UK sample of children and young people aged between four and 16 years old and receiving school-based counselling (n = 6712 students from 308 primary and 61 secondary schools). Fixed period school exclusion rates (number of sessions) were compared between the academic year before and the academic year in which the child attended counselling. Mental health (Strengths and Difficulties Questionnaire) was compared at baseline and at the end of the intervention (after between 16-22 counselling sessions depending on the phase of education).Despite more complex and severe initial difficulties, and facing greater adversity, children and young people who experienced school exclusion prior to counselling demonstrated a significant reduction in subsequent sessions of school exclusion in the academic year that the counselling took place (from two full school weeks to half a school week). Moreover, over 74% of the students had fewer reported exclusions and more than half (56.14%) did not have any further subsequent exclusions. They also had better mental health measured by the teacher reported SDQ (pre-intervention M = 18.94, SD = 6.83 vs. postintervention M = 15.67, SD = 7.56, t(310) = 8.23, p .001) or by the parents (pre-intervention M = 18.09, SD = 6.42 vs. postintervention M = 14.0, SD = 6.99, t(171) = 7.71, p .001).School-based mental health interventions may positively influence educational engagement as well as mental health. Providers should, therefore, monitor both to explore the impact of their interventions. The identification of poor mental health may alter staff perceptions and management of challenging pupils, which future studies should explore.

Background: Poor mental health reduces children’s ability to function at school, which in turn may undermine their mental health. The provision of school-based counselling aims to help schools to support their pupil’s mental health. Most work on the types of difficulties brought to school counsellors has focused on secondary school pupils (aged 11 years or over) and to our knowledge, this is the first study of presenting issues in younger children. Method: Data were systematically collected using a list of 21 potential presenting issues during assessment and formulation of 8,893 children referred for counselling in 291 UK primary schools over 3 years. We explored the number, severity and types of presenting issues recorded by counsellors in the whole sample by gender. Results: The children assessed by counsellors had higher levels of known correlates for poor mental health than their peers in the same schools. Most children had multiple presenting issues, while 55% of children had at least one severe presenting issue. The commonest presenting issues were generalised anxiety, low self-esteem, family tensions and mood swings. Girls were more likely to present with all types of anxiety and family tensions, while attentional problems and mood swings were commoner among boys. Conclusions: Children referred for counselling in primary schools often present with multiple difficulties, which are often severe. This indicates the need for systematic and detailed assessment, adequate training and supervision and good links with external agencies.

The use of systematic client feedback tools are known to enhance outcomes in adults psychotherapy clients, but their effects with children have yet to be adequately tested. Hence, we piloted a clus...

Bacteria have evolved sophisticated adaptive immune systems, called CRISPR-Cas, that provide sequence-specific protection against phage infection. In turn, phages have evolved a broad spectrum of anti-CRISPRs that suppress these immune systems. Here we report structures of anti-CRISPR protein IF9 (AcrIF9) in complex with the type I-F CRISPR RNA-guided surveillance complex (Csy). In addition to sterically blocking the hybridization of complementary dsDNA to the CRISPR RNA, our results show that AcrIF9 binding also promotes non-sequence-specific engagement with dsDNA, potentially sequestering the complex from target DNA. These findings highlight the versatility of anti-CRISPR mechanisms utilized by phages to suppress CRISPR-mediated immune systems., The anti-CRISPR protein IF9 (AcrIF9) specifically inhibits the type I-F CRISPR adaptive immune system. Here, the authors present the cryo-EM structure of AcrIF9 in complex with the type I-F CRISPR RNA-guided surveillance complex (Csy) and a dsDNA bound Csy-AcrIF9 structure, and find that AcrIF9 binding to the Csy complex triggers non-sequence specific dsDNA binding to Csy-AcrIF9, which might sequester the complex from its target DNA.

Bacteria and archaea rely on CRISPR (clustered regularly interspaced short palindromic repeats) RNA-guided adaptive immune systems for sequence specific elimination of foreign nucleic acids. In Escherichia coli, short CRISPR-derived RNAs (crRNAs) assemble with Cas (CRISPR-associated) proteins into a 405-kilodalton multi-subunit surveillance complex called Cascade (CRISPR-associated complex for antiviral defense). Cascade binds foreign DNA complementary to the crRNA guide and recruits Cas3, a trans-acting nuclease-helicase required for target degradation. Structural models of Cascade have captured static snapshots of the complex in distinct conformational states, but conformational dynamics of the 11-subunit surveillance complex have not been measured. Here we use hydrogen-deuterium exchange coupled to mass spectrometry (HDX-MS) to map conformational dynamics of Cascade onto the three-dimensional structure. New insights from structural dynamics are used to make functional predictions about the mechanisms of the R-loop coordination and Cas3 recruitment. We test these predictions in vivo and in vitro. Collectively, we show how mapping conformational dynamics onto static 3D-structures adds an additional dimension to the functional understanding of this biological machine.

Bacteria and archaea have evolved sophisticated adaptive immune systems that rely on CRISPR RNA (crRNA)-guided detection and nuclease-mediated elimination of invading nucleic acids. Here we present the cryo-EM structure of the type I-F CRISPR RNA-guided surveillance complex (Csy complex) from Pseudomonas aeruginosa bound to a double-stranded DNA target. Comparison of this structure to previously determined structures of this complex reveals a Ȉ180-degree rotation of the C-terminal helical bundle on the “large” Cas8f subunit. We show that the dsDNA-induced conformational change in Cas8f exposes a Cas2/3 “nuclease recruitment helix” that is structurally homologous to a virally encoded anti-CRISPR protein (AcrIF3). Structural homology between Cas8f and AcrIF3 suggests that AcrIF3 is a mimic of the Cas8f “nuclease recruitment helix”, implying that cas genes may sometimes serve as genetic fodder for the evolution of anti-CRISPRs.

Summary Bacteria and archaea have evolved sophisticated adaptive immune systems that rely on CRISPR RNA (crRNA)-guided detection and nuclease-mediated elimination of invading nucleic acids. Here, we present the cryo-electron microscopy (cryo-EM) structure of the type I-F crRNA-guided surveillance complex (Csy complex) from Pseudomonas aeruginosa bound to a double-stranded DNA target. Comparison of this structure to previously determined structures of this complex reveals a ∼180-degree rotation of the C-terminal helical bundle on the “large” Cas8f subunit. We show that the double-stranded DNA (dsDNA)-induced conformational change in Cas8f exposes a Cas2/3 “nuclease recruitment helix” that is structurally homologous to a virally encoded anti-CRISPR protein (AcrIF3). Structural homology between Cas8f and AcrIF3 suggests that AcrIF3 is a mimic of the Cas8f nuclease recruitment helix.

In bacteria and archaea, short fragments of foreign DNA are integrated into Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) loci, providing a molecular memory of previous encounters with foreign genetic elements. In Escherichia coli, short CRISPR-derived RNAs are incorporated into a multi-subunit surveillance complex called Cascade (CRISPR-associated complex for antiviral defense). Recent structures of Cascade capture snapshots of this seahorse-shaped RNA-guided surveillance complex before and after binding to a DNA target. Here we determine a 3.2 Å x-ray crystal structure of Cascade in a new crystal form that provides insight into the mechanism of double-stranded DNA binding. Molecular dynamic simulations performed using available structures reveal functional roles for residues in the tail, backbone and belly subunits of Cascade that are critical for binding double-stranded DNA. Structural comparisons are used to make functional predictions and these predictions are tested in vivo and in vitro. Collectively, the results in this study reveal underlying mechanisms involved in target-induced conformational changes and highlight residues important in DNA binding and protospacer adjacent motif recognition.

The type I-F CRISPR adaptive immune system in Pseudomonas aeruginosa (PA14) consists of two CRISPR loci and six CRISPR-associated (cas) genes. Type I-F systems rely on a CRISPR RNA (crRNA)-guided surveillance complex (Csy complex) to bind foreign DNA and recruit a trans-acting nuclease (i.e., Cas2/3) for target degradation. In most type I systems, Cas2 and Cas3 are separate proteins involved in adaptation and interference, respectively. However, in I-F systems, these proteins are fused into a single polypeptide. Here we use biochemical and structural methods to show that two molecules of Cas2/3 assemble with four molecules of Cas1 (Cas2/32:Cas14) into a four-lobed propeller-shaped structure, where the two Cas2 domains form a central hub (twofold axis of symmetry) flanked by two Cas1 lobes and two Cas3 lobes. We show that the Cas1 subunits repress Cas2/3 nuclease activity and that foreign DNA recognition by the Csy complex activates Cas2/3, resulting in bidirectional degradation of DNA targets. Collectively, this work provides a structure of the Cas1–2/3 complex and explains how Cas1 and the target-bound Csy complex play opposing roles in the regulation of Cas2/3 nuclease activity.

Human IBD, including UC and Crohn's disease, is characterized by a chronic, relapsing, and remitting condition that exhibits various features of immunological inflammation and affects at least one/1000 people in Western countries. Polyphenol extracts from a variety of plants have been shown to have immunomodulatory and anti-inflammatory effects. In this study, treatment with APP was investigated to ameliorate chemically induced colitis. Oral but not peritoneal administration of APP during colitis induction significantly protected C57BL/6 mice against disease, as evidenced by the lack of weight loss, colonic inflammation, and shortening of the colon. APP administration dampened the mRNA expression of IL-1β, TNF-α, IL-6, IL-17, IL-22, CXCL9, CXCL10, CXCL11, and IFN-γ in the colons of mice with colitis. APP-mediated protection requires T cells, as protection was abated in Rag-1−/− or TCRα−/− mice but not in IL-10−/−, IRF-1−/−, μMT, or TCRδ−/− mice. Administration of APP during colitis to TCRα−/− mice actually enhanced proinflammatory cytokine expression, further demonstrating a requirement for TCRαβ cells in APP-mediated protection. APP treatment also inhibited CXCR3 expression by TCRαβ cells, but not B or NK cells, in the colons of mice with colitis; however, depletion of CD4+ or CD8+ T cells alone did not abolish APP-mediated protection. Collectively, these results show that oral administration of APP protects against experimental colitis and diminishes proinflammatory cytokine expression via T cells.

Background Protein-energy wasting is common in chronic kidney disease and is associated with decreases in body muscle and fat stores and poor outcomes. The accuracy and reliability of field methods to measure body composition is unknown in this population. Study Design Cross-sectional observational study. Setting & Participants 118 maintenance hemodialysis patients were seen at the General Clinical Research Center at Harbor-UCLA Medical Center, Torrance, CA. Index Tests Triceps skinfold, near-infrared interactance, and bioelectrical impedance analysis using the Segal, Kushner, and Lukaski equations. Reference Test Dual-energy x-ray absorptiometry (DEXA). Results Participants (42% women, 52% with diabetes, 40% African Americans, and 38% Hispanics) were aged 49.4 ± 11.5 (mean ± SD) years, and had undergone dialysis therapy for 41.1 ± 32.9 months. Body mass index was 27.0 ± 6.0 kg/m 2 . Using DEXA as the reference test, the bioelectrical impedance analysis–Kushner equation, triceps skinfold, and near-infrared interactance were most accurate of the index tests in estimating total-body fat percentage, whereas bioelectrical impedance analysis–Segal equation and bioelectrical impedance analysis–Lukaski equation overestimated total body fat percentage. Bland-Altman analyses and difference plots showed that bioelectrical impedance analysis–Kushner and near-infrared interactance were most similar to the reference test. Bioelectrical impedance analysis–Kushner, triceps skinfold, and near-infrared interactance had the smallest mean differences from DEXA, especially in women (1.6%, 0.7%, and 1.2%, respectively). Similar results were observed in African American participants (n = 47). Limitations Measurements were performed 1 day after a hemodialysis treatment, leading to more fluid retention, which may have affected the reference and index tests differently. Conclusions Using DEXA as the reference test, both near-infrared interactance and bioelectrical impedance analysis–Kushner method yield more consistent estimates of total body fat percentage in maintenance hemodialysis patients compared with the other index tests. Near-infrared interactance is not affected by skin color. Field methods with portable devices may provide adequate precision.

The chemokine, lymphotactin (LTN), was tested as a molecular adjuvant using bicistronic DNA vaccines encoding the protective Yersinia capsular (F1) antigen and virulence antigen (V-Ag) as a F1-V fusion protein. The LTN-encoding F1-V or V-Ag vaccines were given by the intranasal (i.n.) or intramuscular (i.m.) routes, and although serum IgG and mucosal IgA antibodies (Abs) were induced, F1-Ag boosts were required for robust anti-F1-Ag Abs. Optimal efficacy against pneumonic plague was obtained in mice i.m.-, not i.n.-immunized with these DNA vaccines. These vaccines stimulated elevated Ag-specific Ab-forming cells and mixed Th cell responses, with Th17 cells markedly enhanced by i.m. immunization. These results show that LTN can be used as a molecular adjuvant to enhance protective immunity against plague., Vaccine, 28(18), pp.3219-3230; 2010

Previous studies have shown that mucosal application of interleukin-12 (IL-12) can stimulate elevated secretory immunoglobulin A (IgA) responses. Since possible exposure to plague is viaYersinia pestis-laden aerosols that results in pneumonic plague, arming both the mucosal and systemic immune systems may offer an added benefit for protective immunity. Two bicistronic plasmids were constructed that encoded the protective plague epitopes, capsular antigen (F1-Ag) and virulence antigen (V-Ag) as a F1-V fusion protein but differed in the amounts of IL-12 produced. When applied nasally, serum IgG and mucosal IgA anti-F1-Ag and anti-V-Ag titers were detectable beginning at week 6 after three weekly doses, and recombinant F1-Ag boosts were required to elevate the F1-Ag-specific antibody (Ab) titers. Following pneumonic challenge, the best efficacy was obtained in mice primed with IL-12(Low)/F1-V vaccine with 80% survival compared to mice immunized with IL-12(Low)/F1, IL-12(Low)/V, or IL-12(Low) vector DNA vaccines. Improved expression of IL-12 resulted in lost efficacy when using the IL-12(High)/F1-V DNA vaccine. Despite differences in the amount of IL-12 produced by the two F1-V DNA vaccines, Ab responses and Th cell responses to F1- and V-Ags were similar. These results show that IL-12 can be used as a molecular adjuvant to enhance protective immunity against pneumonic plague, but in a dose-dependent fashion.

Genetic conflict between viruses and their hosts drives evolution and genetic innovation. Prokaryotes evolved CRISPR (Clustered Regularly Interspaced Short Palindromic Repeat)-mediated adaptive immune systems for protection from viral infection and viruses have evolved diverse anti-CRISPR (Acr) proteins that subvert these immune systems. The adaptive immune system in Pseudomonas aeruginosa (type I-F) relies on a 350 kDa CRISPR RNA (crRNA)-guided surveillance complex (Csy complex) to bind foreign DNA and recruit a trans-acting nuclease for target degradation. Here we report the cryo-electron microscopy structure of the Csy complex bound to two different Acr proteins, AcrF1 and AcrF2, at an average resolution of 3.4 Å. The structure explains the molecular mechanism for immune system suppression, and structure-guided mutations show that the Acr proteins bind to residues essential for crRNA-mediated detection of DNA. Collectively, these data provide a snapshot of an ongoing molecular arms race between viral suppressors and the immune system they target.

A foreign-DNA–destroying machine Bacteria have an adaptive immune system, called CRISPR, that identifies invading viruses through their DNA or RNA sequences and cuts them up (see the Perspective by Zhang and Sontheimer). Jackson et al. and Mulepati et al. have determined the structure of the large protein complex, called Cascade, that targets the invading nucleic acids and does the cutting. The seahorse-shaped structure reveals how the 11 subcomponents of Cascade assemble into the final protein complex. The structure also shows how Cascade presents the short CRISPR-derived RNAs so that they can bind and target foreign DNA. Science , this issue p. 1473 and p. 1479 ; see also p. 1452

Our paper focuses on the environmental disclosures made by 51 United States (US) manufacturing firms for 1992 and 1993. Content analysis of annual report disclosures, made within the president's letter, the management's discussion and analysis (MD&A), and note sections, was used in conjunction with environmental ratings of US corporations, as compiled by the Council on Economic Priorities (CEP). Our study investigated whether disclosures differed between firms who had been rated good, mixed or poor in their environmental activities and whether these disclosure differences could be used to differentiate between actual environmental performance levels. Our study also investigated how a change in disclosure standard for one section of the annual report changes disclosures in other sections of the annual report. The results are important in light of Dunlap and Scarce's (1991, p. 657) findings that public concern about the environment was at an all-time high, and Epstein and Freedman's (1994, p. 106) finding that individual investors consider annual report information about environmental activities to be more desirable than information about any other social activity.

Background: We tested the hypothesis that cycle length-dependent cardiac contractility in atrial fibrillation is primarily governed by the negative interval-force relation in patients with normal and depressed systolic function. Methods and Results: We performed two-dimensional guided M-mode echocardiography in 41 patients (mean age, 69 ± 4 years; range, 48 to 92 years; 19 men, 11 women). Twelve patients had objective evidence of left ventricular systolic dysfunction (CMP; mean ejection fraction, 37% ± 7%) in the absence of coronary artery disease (CAD), 13 patients had documented CAD (mean ejection fraction, 43% ± 6%), and 16 patients had normal resting left ventricular systolic function (mean ejection fraction, 58% ± 7%). Simultaneous beat-to-beat blood pressure, end-systolic and end-diastolic dimension, circumferential velocity of fiber shortening (Vcf), and end-systolic wall stress (ESWS) were calculated for all patients. All three groups showed a significant linear relation between beat-to-beat Vcf and Vcf corrected for afterload (represented as the Vcf/ESWS ratio) and preceding cycle length. There was, however, no significant difference in the relation between either of these variables and cycle length among the three groups. There was also no difference in the rate of change in either Vcf or Vcf corrected for afterload (Vcf/ESWS ratio) from beat-to-beat among the three groups. Control patients with normal systolic function showed greater Vcf at any given cycle length compared with patients with CMP or CAD. Conclusion: Our data show that, for each beat in atrial fibrillation, Vcf and Vcf/ESWS ratio are decreased after shorter cycle lengths and increased after long cycles, but there is no significant attenuation of this effect in patients with systolic dysfunction with or without coronary disease compared with controls. Thus, the negative interval-force relation, the predominant determinant of beat-to-beat variation in contractility in atrial fibrillation, is preserved in patients with CAD or reduced left ventricular systolic function.

In recent years, mass participation in post-16 education and training in England has led to a diminishing understanding about young people who leave education at the end of compulsory schooling to enter ‘jobs without training’ (JWT). Drawing on data from three recent studies, this article argues that the JWT group is not homogeneous in its composition. Similar findings led to the development of a common typology across all three studies to define young people's position in the labour market, their motivations and aspirations, and their access to training and development. It concludes with a series of recommendations for addressing the deficit in knowledge about the composition of the JWT group, and the learning and training needs of young workers. This discussion is set in the context of the implementation of the Raising of the Participation Age (RPA) in England for all 17-year olds from 2013 and for all 18-year olds from 2015, although within the Coalition Government's current proposals, its delivery will lack any form of immediate enforcement. Therefore, unless young workers and their employers are committed to the acquisition of accredited qualifications, RPA delivery will be seriously undermined and intervention to support school to work transitions among the JWT group will remain negligible.

Osteoarticular and soft tissue inflammation occur in subsequent infection with wild-type Brucella in mice deficient of IFN-γ, resembling aspects of human disease. Human brucellosis exhibits diverse pathological manifestations that can affect almost any organ. In particular, osteoarticular complications are the most common focal manifestation of brucellosis and occur in 40–80% of patients. In immunocompetent mice, Brucella replication is generally restricted to the spleen, liver, and to a lesser extent, LNs, thereby limiting their use for study of focal inflammation often found in brucellosis. Here, we report that nasal, oral, or peritoneal infection of IFN-γ−/− mice with WT Brucella melitensis or Brucella abortus results in joint and periarticular tissue inflammation. Histological analysis of the affected joints revealed inflammatory infiltrates and debris within the joint space colocalizing with Brucella antigen. Osteoarthritis, necrosis, periarticular soft tissue inflammation, and substantial brucellae burdens were observed. Oral rifampicin was effective in clearing infection and halting further progression of focal inflammation from infected IFN-γ−/− mice, although some symptoms and swelling remained. Elevated IL-1β, but not TNF-α, IL-6, or IL-17, was detected in joint homogenates from infected IFN-γ−/− mice. Whereas more susceptible to systemic infection, IL-1R−/− mice depleted of IFN-γ were more resistant to focal inflammation than WT mice similarly depleted of IFN-γ. Collectively, these results show IFN-γ−/− mice represent a potential model for study of focal inflammation attributed to Brucella infection and will allow evaluation of intervention strategies targeting IL-1, IL-1R, or other inflammatory mediators, with the potential to complement antibiotic-based therapies.

Natural killer (NK) cells and dendritic cells (DCs) have been shown to link the innate and adaptive immune systems. Likewise, a new innate cell subset, interferon-producing killer DCs (IKDCs), shares phenotypic and functional characteristics with both DCs and NK cells. Here, we show IKDCs play an essential role in the resolution of experimental autoimmune encephalomyelitis (EAE) upon treatment with the tolerizing agent, myelin oligodendrocyte glycoprotein (MOG), genetically fused to reovirus protein σ1 (termed MOG-pσ1). Activated IKDCs were recruited subsequent MOG-pσ1 treatment of EAE, and disease resolution was abated upon NK1.1 cell depletion. These IKDCs were able to kill activated CD4(+) T cells and mature dendritic DCs, thus, contributing to EAE remission. In addition, IKDCs were responsible for MOG-pσ1-mediated MOG-specific regulatory T cell recruitment to the CNS. The IKDCs induced by MOG-pσ1 expressed elevated levels of HVEM for interactions with cognate ligand-positive cells: LIGHT(+) NK and T(eff) cells and BTLA(+) B cells. Further characterization revealed these activated IKDCs being MHC class II(high), and upon their adoptive transfer (CD11c(+)NK1.1(+)MHC class II(high)), IKDCs, but not CD11c(+)NK1.1(+)MHC class II(intermediate/low) (unactivated) cells, conferred protection against EAE. These activated IKDCs showed enhanced CD107a, PD-L1, and granzyme B expression and could present OVA, unlike unactivated IKDCs. Thus, these results demonstrate the interventional potency induced HVEM(+) IKDCs to resolve autoimmune disease.

Background Immunization with recombinant carboxyl-terminal domain of the heavy chain (Hc domain) of botulinum neurotoxin (BoNT) stimulates protective immunity against native BoNT challenge. Most studies developing a botulism vaccine have focused on the whole Hc; however, since the principal protective epitopes are located within β-trefoil domain (Hcβtre), we hypothesize that immunization with the Hcβtre domain is sufficient to confer protective immunity. In addition, enhancing its uptake subsequent to nasal delivery prompted development of an alternative vaccine strategy, and we hypothesize that the addition of targeting moiety adenovirus 2 fiber protein (Ad2F) may enhance such uptake during vaccination. Results The Hcβtre serotype B immunogen was genetically fused to Ad2F (Hcβtre/B-Ad2F), and its immunogenicity was tested in mice. In combination with the mucosal adjuvant, cholera toxin (CT), enhanced mucosal IgA and serum IgG Ab titers were induced by nasal Hcβtre-Ad2F relative to Hcβtre alone; however, similar Ab titers were obtained upon intramuscular immunization. These BoNT/B-specific Abs induced by nasal immunization were generally supported in large part by Th2 cells, as opposed to Hcβtre-immunized mice that showed more mixed Th1 and Th2 cells. Using a mouse neutralization assay, sera from animals immunized with Hcβtre and Hcβtre-Ad2F protected mice against 2.0 LD50. Conclusion These results demonstrate that Hcβtre-based immunogens are highly immunogenic, especially when genetically fused to Ad2F, and Ad2F can be exploited as a vaccine delivery platform to the mucosa.

IL-35 is produced by regulatory T cells, and this novel cytokine can downregulate Th17 cell development and inhibit autoimmune inflammation. In this work, an rIL-35, as a single-chain fusion between murine IL-12p35 and EBV-induced gene 3, was expressed in yeast. This rIL-35 inhibited OVA-specific cellular and Ab responses in OVA-challenged recipients of DO11.10 CD4+ T cells. Likewise, IL-35 inhibited clinical manifestation of collagen-induced arthritis or could cease further disease exacerbation upon initiation of IL-35 treatment. Exogenous IL-35 treatments suppressed Th1 and Th17 cells and promoted CD39 expression by CD4+ T cells. Sorted CD25−CD39+CD4+ T cells from IL-35–treated mice produced IL-10 and, upon adoptive transfer, were sufficiently potent to inhibit subsequent development of inflammation in mice with collagen-induced arthritis, whereas sorted CD25+CD39+CD4+ T cells showed reduced potency. IL-35 treatments of IL-10−/− mice failed to induce protective CD39+CD4+ T cells, demonstrating the effector role of IL-10 by IL-35 immunosuppression.

The psychological factors that predispose an individual to substance abuse are the same factors involved in relapse, a major threat to recovery from a reliance on substances. Modified Dynamic Group Therapy (MDGT) is a group therapeutic approach that is sufficiently supportive and structured to provide the safety and comfort that such patients require, but unstructured enough to allow a natural unfolding of their vulnerabilities and characterological defenses that need to be identified and modified. MDGT targets and focuses on four areas of self-regulation vulnerabilities involving difficulties with feeling life (affects), self-esteem, relationships, and self-care, and the defensive, charactemlogical traits that compensate for, while they also reveal, these vulnerabilties.

The psychological factors that predispose an individual to substance abuse are the same factors involved in relapse, a major threat to recovery from a reliance on substances. Modified Dynamic Group Therapy (MDGT) is a group therapeutic approach that is sufficiently supportive and structured to provide the safety and comfort that such patients require, but unstructured enough to allow a natural unfolding of their vulnerabilities and characterological defenses that need to be identified and modified. MDGT targets and focuses on four areas of self-regulation vulnerabilities involving difficulties with feeling life (affects), self-esteem, relationships, and self-care, and the defensive, characterological traits that compensate for, while they also reveal, these vulnerabilties.

Pneumonic plague remains problematic in endemic areas, and because it can be readily transmitted and has high mortality, the development of efficacious vaccines is warranted. To test whether stimulation of cell-mediated immunity with IL-12 will improve protective immunity against plague, we constructed two IL-12 DNA vaccines using a bicistronic plasmid encoding the protective plague epitopes, capsular (F1) antigen and virulence antigen (V-Ag) as F1-V fusion protein and V-Ag only, respectively. When applied intramuscularly, antibody responses to F1- and V-Ag were detectable beginning at week 6 after three weekly doses, and F1-Ag protein boosts were required to induce elevated Ab responses. These Ab responses were supported by mixed Th cell responses, and the IL-12/V-Ag DNA vaccine showed greater cell-mediated immune bias than IL-12/F1-V DNA vaccine. Following pneumonic challenge, both IL-12 DNA vaccines showed similar efficacy despite differences in Th cells simulated. These results show that IL-12 can be used as a molecular adjuvant to enhance protective immunity against pneumonic plague.

Clinicians providing psychoeducation give information and support to family members in a nonjudgmental, educational format (1). To improve the ftmily’s understanding of their relative’s overall treatment, psychoeducation embraces evolving biological theories of mental illness and promotes a sense of “cognitive mastery” (1) of the family member’s illness and its management. The efficacy of the psychoeducational approach has been supported by early research demonstrating that it decreased on forestalled patients’ relapse and nehospitalization (1 ,2). In recent years, the theory of expressed emotion has been formulated, which suggests that psychoeducation improves outcome by decreasing the family’s emotional intensity and creating a more benign attitude toward the patient (3). This paper is based on six cliicians’ collective experience doing psychoeducational group work with

Mucosal intervention with CFA/I fimbriae effectively diminishes CIA by stimulation of TGF-β+ and IL-10+ regulatory CD39+CD4+ T cells (Tregs), resulting in suppression of Th1 and Th17 cells and IL-27. CFA/I also stimulates Tregs expressing p35 and EBI3 subunits simultaneously, suggesting an essential role of IL-35 in CFA/I-mediated protection. We hypothesize CFA/I fimbriae stimulate endogenous IL-35 production to facilitate protection against autoimmune disease. In testing the relevance of endogenous IL-35 for protection, oral delivery of recombinant CFA/I into EBI3-/- mice on day 14 after CIA induction was found ineffective against CIA compared to similarly treated WT mice, as evidenced by the lack of suppression of IL-17 and IFN-γ (the latter increased 5-fold ). CFA/I-treated EBI3-/- mice lacked enhanced CREB phosphorylation that, in turn, failed to stimulate the transcription of CD39 (apyrase). In fact, surface apyrase activity by EBI3-/- CD4+ T cells was uninduced. Because of the lack of induced CD39 expression, IL-10 production was greatly diminished. Clinical improvement of CIA in CFA/I-treated EBI3-/- mice required further intervention with recombinant IL-35, resulting in the induction of CD39+CD4+ T cells, enhanced TGF-β production, and reduction in IFN-γ and IL-17. Thus, in the absence of endogenous IL-35, mucosal CFA/I fimbriae intervention is ineffective against autoimmune arthritis. Supported by AT-004986.

Infection of the joints is the most frequent localized manifestation of brucellosis, which is a common cause of infectious arthritis. However, no experimental murine model of Brucella-induced arthritis has been reported. Here we report that IFN-γ-/- mice develop joint inflammation following oral, nasal, or parenteral infection with B. abortus or B. melitensis. Joints from Brucella-infected IFN-γ-/-, but not wild-type mice, were found to contain extensive inflammatory infiltrates and debris within the joint space which co-localized with brucellae. Osteoarthritis, joint space narrowing, necrosis, soft tissue inflammation, and substantial Brucella burdens were also observed, although antibody or cytokine responses against collagen were not detected. Elevated IL-1β, but not TNF-α, IL-6, nor IL-17, was detected in the joints of Brucella-infected IFN-γ-/- mice. A six-week regimen of rifampicin effectively cleared infection and halted further progression of inflammation, although some symptoms and swelling remained. However, administration of an adenovirus expressing the IL-1receptor antagonist augmented antibiotic resolution of joint swelling by >50%. These results show that the IFN-γ-/- mouse represents a useful model to study the pathogenesis of joint inflammation due to Brucella infection, and that intervention strategies targeting IL-1 can complement antibiotic treatment of Brucella-induced inflammation. Supported by USDA 2007-01612 and USDA 2009-34397-20133.

IL-35, a novel heterodimeric cytokine (IL-12p35 subunit plus Epstein-Barr virus-induced gene 3 [EBI3]), had been reported to expand regulatory CD4+CD25+ T cells and ameliorate CIA in mice. IL-35 was expressed as a single chain polypeptide, and the recombinant cytokine was detected by Western blot using anti-EBI3 or anti-IL-12 mAbs. To assess its activity, BALB/c mice were adoptively transferred with DO11.10 CD4+ T cells, challenged s.c. with OVA in IFA, and treated every day for 5 days with IL-35, resulting in diminished IgG2a and IgG2b anti-OVA Abs, as well as suppressed DTH responses. To exploit IL-35's therapeutic potential, B6 mice were induced with CIA, and upon the onset of disease (day 21), mice received 2.0 μg/mouse every 2 days until day 39. All mice in the control group developed arthritis by day 28 post-induction, whereas none of IL-35-treated mice had any clinical symptoms. Restimulated sorted CD4+ T cells from LNs of protected mice showed decreased IFN-γ and IL-17 production. FACS analysis revealed enhancement of regulatory CD4+ CD25+ T cells, of which, half co-expressed CD39, an ectonucleoside triphosphate diphosphohydrolase. Cell-sorted CD39+ CD4+ T cells from IL-35-treated mice could suppress CII-specific proliferation by CD39- CD4+ T cells via enhanced IL-10 production. These findings suggest the stimulation by IL-35 for the induction of regulatory CD4+ T cells. Supported by NIH R01 AI78938.