Clinical effectiveness, cost-effectiveness and acceptability of low-intensity interventions in the management of obsessive-compulsive disorder: the Obsessive-Compulsive Treatment Efficacy randomised controlled Trial (OCTET).

BACKGROUND: The Obsessive-Compulsive Treatment Efficacy randomised controlled Trial emerged from a research recommendation in National Institute for Health and Care Excellence obsessive-compulsive disorder (OCD) guidelines, which specified the need to evaluate cognitive-behavioural therapy (CBT) treatment intensity formats. OBJECTIVES: To determine the clinical effectiveness and cost-effectiveness of two low-intensity CBT interventions [supported computerised cognitive-behavioural therapy (cCBT) and guided self-help]: (1) compared with waiting list for high-intensity CBT in adults with OCD at 3 months; and (2) plus high-intensity CBT compared with waiting list plus high-intensity CBT in adults with OCD at 12 months. To determine patient and professional acceptability of low-intensity CBT interventions. DESIGN: A three-arm, multicentre, randomised controlled trial. SETTING: Improving Access to Psychological Therapies services and primary/secondary care mental health services in 15 NHS trusts. PARTICIPANTS: Patients aged ≥ 18 years meeting Diagnostic and Statistical Manual of Mental Disorders-Fourth Edition criteria for OCD, on a waiting list for high-intensity CBT and scoring ≥ 16 on the Yale-Brown Obsessive Compulsive Scale (indicative of at least moderate severity OCD) and able to read English. INTERVENTIONS: Participants were randomised to (1) supported cCBT, (2) guided self-help or (3) a waiting list for high-intensity CBT. MAIN OUTCOME MEASURES: The primary outcome was OCD symptoms using the Yale-Brown Obsessive Compulsive Scale - Observer Rated. RESULTS: Patients were recruited from 14 NHS trusts between February 2011 and May 2014. Follow-up data collection was complete by May 2015. There were 475 patients randomised: supported cCBT (n = 158); guided self-help (n = 158) and waiting list for high-intensity CBT (n = 159). Two patients were excluded post randomisation (one supported cCBT and one waiting list for high-intensity CBT); therefore, data were analysed for 473 patients. In the short term, prior to accessing high-intensity CBT, guided self-help demonstrated statistically significant benefits over waiting list, but these benefits did not meet the prespecified criterion for clinical significance [adjusted mean difference -1.91, 95% confidence interval (CI) -3.27 to -0.55; p = 0.006]. Supported cCBT did not demonstrate any significant benefit (adjusted mean difference -0.71, 95% CI -2.12 to 0.70). In the longer term, access to guided self-help and supported cCBT, prior to high-intensity CBT, did not lead to differences in outcomes compared with access to high-intensity CBT alone. Access to guided self-help and supported cCBT led to significant reductions in the uptake of high-intensity CBT; this did not seem to compromise patient outcomes at 12 months. Taking a decision-making approach, which focuses on which decision has a higher probability of being cost-effective, rather than the statistical significance of the results, there was little evidence that supported cCBT and guided self-help are cost-effective at the 3-month follow-up compared with a waiting list. However, by the 12-month follow-up, data suggested a greater probability of guided self-help being cost-effective than a waiting list from the health- and social-care perspective (60%) and the societal perspective (80%), and of supported cCBT being cost-effective compared with a waiting list from both perspectives (70%). Qualitative interviews found that guided self-help was more acceptable to patients than supported cCBT. Professionals acknowledged the advantages of low intensity interventions at a population level. No adverse events occurred during the trial that were deemed to be suspected or unexpected serious events. LIMITATIONS: A significant issue in the interpretation of the results concerns the high level of access to high-intensity CBT during the waiting list period. CONCLUSIONS: Although low-intensity interventions are not associated with clinically significant improvements in OCD symptoms, economic analysis over 12 months suggests that low-intensity interventions are cost-effective and may have an important role in OCD care pathways. Further research to enhance the clinical effectiveness of these interventions may be warranted, alongside research on how best to incorporate them into care pathways. TRIAL REGISTRATION: Current Controlled Trials ISRCTN73535163. FUNDING: This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 21, No. 37. See the NIHR Journals Library website for further project information.

Low-intensity cognitive-behaviour therapy interventions for obsessive-compulsive disorder compared to waiting list for therapist-led cognitive-behaviour therapy: 3-arm randomised controlled trial of clinical effectiveness.

BACKGROUND: Obsessive-compulsive disorder (OCD) is prevalent and without adequate treatment usually follows a chronic course. "High-intensity" cognitive-behaviour therapy (CBT) from a specialist therapist is current "best practice." However, access is difficult because of limited numbers of therapists and because of the disabling effects of OCD symptoms. There is a potential role for "low-intensity" interventions as part of a stepped care model. Low-intensity interventions (written or web-based materials with limited therapist support) can be provided remotely, which has the potential to increase access. However, current evidence concerning low-intensity interventions is insufficient. We aimed to determine the clinical effectiveness of 2 forms of low-intensity CBT prior to high-intensity CBT, in adults meeting the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV) criteria for OCD. METHODS AND FINDINGS: This study was approved by the National Research Ethics Service Committee North West-Lancaster (reference number 11/NW/0276). All participants provided informed consent to take part in the trial. We conducted a 3-arm, multicentre randomised controlled trial in primary- and secondary-care United Kingdom mental health services. All patients were on a waiting list for therapist-led CBT (treatment as usual). Four hundred and seventy-three eligible patients were recruited and randomised. Patients had a median age of 33 years, and 60% were female. The majority were experiencing severe OCD. Patients received 1 of 2 low-intensity interventions: computerised CBT (cCBT; web-based CBT materials and limited telephone support) through "OCFighter" or guided self-help (written CBT materials with limited telephone or face-to-face support). Primary comparisons concerned OCD symptoms, measured using the Yale-Brown Obsessive Compulsive Scale-Observer-Rated (Y-BOCS-OR) at 3, 6, and 12 months. Secondary outcomes included health-related quality of life, depression, anxiety, and functioning. At 3 months, guided self-help demonstrated modest benefits over the waiting list in reducing OCD symptoms (adjusted mean difference = -1.91, 95% CI -3.27 to -0.55). These effects did not reach a prespecified level of "clinically significant benefit." cCBT did not demonstrate significant benefit (adjusted mean difference = -0.71, 95% CI -2.12 to 0.70). At 12 months, neither guided self-help nor cCBT led to differences in OCD symptoms. Early access to low-intensity interventions led to significant reductions in uptake of high-intensity CBT over 12 months; 86% of the patients allocated to the waiting list for high-intensity CBT started treatment by the end of the trial, compared to 62% in supported cCBT and 57% in guided self-help. These reductions did not compromise longer-term patient outcomes. Data suggested small differences in satisfaction at 3 months, with patients more satisfied with guided self-help than supported cCBT. A significant issue in the interpretation of the results concerns the level of access to high-intensity CBT before the primary outcome assessment. CONCLUSIONS: We have demonstrated that providing low-intensity interventions does not lead to clinically significant benefits but may reduce uptake of therapist-led CBT. TRIAL REGISTRATION: International Standard Randomized Controlled Trial Number (ISRCTN) Registry ISRCTN73535163.

Obsessive Compulsive Treatment Efficacy Trial (OCTET) comparing the clinical and cost effectiveness of self-managed therapies: study protocol for a randomised controlled trial.

BACKGROUND: UK National Institute of Health and Clinical Excellence guidelines for obsessive compulsive disorder (OCD) specify recommendations for the treatment and management of OCD using a stepped care approach. Steps three to six of this model recommend treatment options for people with OCD that range from low-intensity guided self-help (GSH) to more intensive psychological and pharmacological interventions. Cognitive behavioural therapy (CBT), including exposure and response prevention, is the recommended psychological treatment. However, whilst there is some preliminary evidence that self-managed therapy packages for OCD can be effective, a more robust evidence base of their clinical and cost effectiveness and acceptability is required. METHODS/DESIGN: Our proposed study will test two different self-help treatments for OCD: 1) computerised CBT (cCBT) using OCFighter, an internet-delivered OCD treatment package; and 2) GSH using a book. Both treatments will be accompanied by email or telephone support from a mental health professional. We will evaluate the effectiveness, cost and patient and health professional acceptability of the treatments. DISCUSSION: This study will provide more robust evidence of efficacy, cost effectiveness and acceptability of self-help treatments for OCD. If cCBT and/or GSH prove effective, it will provide additional, more accessible treatment options for people with OCD. TRIAL REGISTRATION: Current Controlled Trials: ISRCTN73535163. Date of registration: 5 April 2011.

Characterisation of local immune responses induced by a novel nano-particle based carrier-adjuvant in sheep.

Most adjuvants require danger signals to promote immune responses against vaccine antigens. Our previous studies have characterised a powerful nano-particulate antigen delivery system, which by itself does not induce inflammation, and which further appears to induce substantial immune responses in mice and sheep without the requirement for added stimulators of toll like receptors or other pathogen recognition receptors. In the present study we dissect the nature of the early induction phase of the immune response stimulated by such a vaccine comprising 40 nm polystyrene nano-particles conjugated to the antigen. We analyse the kinetics of export from an individual draining lymph node from the sheep, of antibodies and cytokines as well as antigen responsive CD4 and CD8 T cells. Our results indicate that simple inert nano-bead based antigen delivery into the draining area of the lymph node is highly efficient at priming combined humoral and T cell antigen specific immunity without the need for added 'danger signals'. Furthermore this nano-bead adjuvant is a potent agent capable of promoting cross-priming for CD8 T cell induction in sheep. Interestingly, using nano-beads, similarly to what has been observed with natural pathogen based lymph node stimulation, a phase of CD4 T cell priming and export preceded CD8 T cell induction, suggesting the engagement of natural priming processes and kinetics.

Transgenic mice expressing human FcgammaRIIa have enhanced sensitivity to induced autoimmune arthritis as well as elevated Th17 cells.

The major human Fc receptor, huFcgammaRIIa, is implicated in the development of autoimmune arthritis in humans but until recently has not been studied in mouse models. We evaluated potential roles of FcgammaRIIa by using transgenic mice expressing the receptor. We examined two models of induced autoimmune arthritis pristane-induced arthritis (PIA) and collagen-induced arthritis (CIA) as well as the anti-collagen-II antibody-induced arthritis (CAIA) model. In the induced arthritis models PIA and CIA, the transgenic mice developed a more severe arthritis than the other arthritis-prone SJL or DBA1 mice. Interestingly, anti-collagen-II antibodies were elevated in PIA in the susceptible mice. In the CIA model, the highly susceptible transgenic mouse had IgG subclass levels equivalent to the unaffected and disease resistant C57BL/6 mouse strain implying that the FcgammaRIIa lowers the threshold of IgG dependent leukocyte activation. This is consistent with the greatly enhanced sensitivity of the FcgammaRIIa transgenic mice to CAIA which clearly indicates a role for the receptor at least at the inflammatory effector cell level. Other roles for huFcgammaRIIa or other gene products in the development of autoimmunity cannot be ruled out however, especially as the mice exhibited elevated Th1 or Th17 CD4 T cells in the draining lymph nodes.

Inhibition of destructive autoimmune arthritis in FcgammaRIIa transgenic mice by small chemical entities.

The interaction of immune complexes with the human Fc receptor, FcgammaRIIa, initiates the release of inflammatory mediators and is implicated in the pathogenesis of human autoimmune diseases, including rheumatoid arthritis and systemic lupus erythematosus, so this FcR is a potential target for therapy. We have used the three-dimensional structure of an FcgammaRIIa dimer to design small molecule inhibitors, modeled on a distinct groove and pocket created by receptor dimerization, adjacent to the ligand-binding sites. These small chemical entities (SCEs) blocked immune complex-induced platelet activation and aggregation and tumor necrosis factor secretion from macrophages in a human cell line and transgenic mouse macrophages. The SCE appeared specific for FcgammaRIIa, as they inhibited only immune complex-induced responses and had no effect on responses to stimuli unrelated to FcR, for example platelet stimulation with arachidonic acid. In vivo testing of the SCE in FcgammaRIIa transgenic mice showed that they inhibited the development and stopped the progression of collagen-induced arthritis (CIA). The SCEs were more potent than methotrexate and anti-CD3 in sustained suppression of CIA. Thus, in vitro and in vivo activity of these SCE FcgammaRIIa receptor antagonists demonstrated their potential as anti-inflammatory agents for autoimmune diseases involving immune complexes.

Middle-ear disease and schizophrenia: case-control study.

BACKGROUND: One hundred years ago psychiatrists thought that ear disease could cause insanity by irritation of the brain. Current understanding of the role of the temporal lobes in schizophrenia and their proximity to the middle ear supports this hypothesis. AIMS: To establish the rate of middle-ear disease pre-dating the onset of schizophrenia. METHOD: Eighty-four patients with schizophrenia were each matched to four non-psychiatric controls by age, gender and season of birth. History of ear disease was obtained from general practice records. Additional information on symptoms was collected for participants in the case group, who also had audiometry. RESULTS: The odds ratio of recorded middle-ear disease pre-dating schizophrenia was 3.68 (95% CI 1.86-7.28). This excess was particularly marked on the left (OR=4.15, 95% CI 2.08-8.29). Auditory hallucinations were associated with middle-ear disease but not with hearing loss. CONCLUSIONS: There is an association between middle-ear disease and schizophrenia which may have aetiological significance.

The dendritic cell subtype-restricted C-type lectin Clec9A is a target for vaccine enhancement.

A novel dendritic cell (DC)-restricted molecule, Clec9A, was identified by gene expression profiling of mouse DC subtypes. Based on sequence similarity, a human ortholog was identified. Clec9A encodes a type II membrane protein with a single extracellular C-type lectin domain. Both the mouse Clec9A and human CLEC9A were cloned and expressed, and monoclonal antibodies (mAbs) against each were generated. Surface staining revealed that Clec9A was selective for mouse DCs and was restricted to the CD8(+) conventional DC and plasmacytoid DC subtypes. A subset of human blood DCs also expressed CLEC9A. A single injection of mice with a mAb against Clec9A, which targets antigens (Ags) to the DCs, produced a striking enhancement of antibody responses in the absence of added adjuvants or danger signals, even in mice lacking Toll-like receptor signaling pathways. Such targeting also enhanced CD4 and CD8 T-cell responses. Thus, Clec9A serves as a new marker to distinguish subtypes of both mouse and human DCs. Furthermore, targeting Ags to DCs with antibodies to Clec9A is a promising strategy to enhance the efficiency of vaccines, even in the absence of adjuvants.

Mental health diagnosis by nurses using the Global Mental Health Assessment Tool: a validity and feasibility study.

BACKGROUND: The Global Mental Health Assessment Tool-Primary Care Version (GMHAT/PC) has been developed to assist health professionals to make a quick and comprehensive standardised mental health assessment. It has proved to be a reliable and valid tool in a previous study involving GPs. Its use by other health professionals may help in detecting and managing mental disorders in primary care and general health settings. AIM: To assess the feasibility of using a computer-assisted diagnostic interview by nurses and to examine the level of agreement between the GMHAT/PC diagnosis and psychiatrists' clinical diagnosis. DESIGN OF STUDY: Cross-sectional validation study. SETTING: Primary care, general healthcare (cardiac rehabilitation clinic), and community mental healthcare settings. METHOD: A total of 215 patients between the ages of 16 and 75 years were assessed by nurses and psychiatrists in various settings: primary care centre (n = 54), cardiac rehabilitation centre (n = 98), and community mental health clinic (n = 63). The time taken for the interview, and feedback from patients and interviewers were indicators of feasibility, and the kappa coefficient (kappa), sensitivity, and specificity of the GMHAT/PC diagnosis were measures of validity. RESULTS: Mean duration of interview was under 15 minutes. The agreement between nurses' GMHAT/PC interview-based diagnosis and psychiatrists' International Classification of Diseases (ICD)-10 criteria-based clinical diagnosis was 80% (kappa = 0.76, sensitivity = 0.84, specificity = 0.92). CONCLUSION: The GMHAT/PC can assist nurses to make accurate mental health assessment and diagnosis in various healthcare settings and it is acceptable to patients.

Survival in the community of the very old depressed, discharged from medical inpatient care.

OBJECTIVES: To examine the prevalence and associated risk factors of depression in older patients discharged home from acute medical care and their influence on duration of survival in the community. DESIGN: A cross-sectional, prevalence study of depression in recently discharged patients and a prospective, case-controlled study of depressed and psychiatrically asymptomatic sub groups, exploring the relationship between depression, associated risk factors, and duration of survival in the community. SETTING: A community study of patients aged 75 and older discharged from the Countess of Chester Hospital and Wirral Hospitals Trust serving Wirral and West Cheshire, England. PARTICIPANTS: Three hundred and eleven patients were entered into the prevalence study. One hundred and fifty-eight patients (54 depressed and 104 asymptomatic) were entered into the prospective case controlled study and followed up for up to two years. MEASUREMENTS: Depression was defined by GMS/AGECAT criteria. Demographic details, handicap, pain, forced expiratory volume and social network were measured as dependent variables in the prevalence study and included in the analysis of risk factors potentially associated with duration of survival in the community. RESULTS: A depression prevalence rate of 17.4% was found. Age (p = 0.049, CI; 0.813, 0.999), forced expiratory volume (p = 0.034, CI; 0.991, 1.000) and handicap (p = 0.000, CI; 1.268, 1.723) were associated with depression but depression (p = 0.040, CI; 1.039, 4.915) was the only base-line variable associated with reduced survival in the community as defined by mortality and re-admission. CONCLUSIONS: Depression is common in older people discharged from acute medical care and is a major risk factor for reduced duration of community survival.

Type 1 and 2 immunity following vaccination is influenced by nanoparticle size: formulation of a model vaccine for respiratory syncytial virus.

Previous studies compared uptake by dendritic cells (DC) of 20, 40, 100, 200, 500, 1000, and 2000 nm beads in vivo. When beads were used as antigen carriers, bead size influenced antibody responses and induction of IFN-gamma-producing CD4 and CD8 T cells. Beads of 40-50 nm were taken up preferentially by DC and induced particularly strong immunity. Herein, we examine immunity induced by minute differences in nanobead size, specifically within a narrow viral-sized range (20, 40, 49, 67, 93, 101, and 123 nm), to see if bead carrier size influenced the induction of type 1 or type 2 cells as demonstrated by the production of IFN-gamma or IL-4. In vivo uptake by DC was assessed for selected sizes in this range. Responses to whole ovalbumin (OVA) or the OVA-derived CD8 T cell peptide epitope (SIINFEKL) were tested. After one immunization with beads-OVA, IFN-gamma responses to both OVA and SIINFEKL were significantly better with 40 and 49 nm beads than other sizes, while, in contrast, IL-4 responses to OVA were higher after immunization with OVA conjugated to larger beads (93, 101, and 123 nm). Thus IFN-gamma induction from CD8 T cells was limited to 40-49 nm beads, while CD4 T cell activation and IL-4 were induced by 93-123 nm beads-OVA. After two immunizations, there were comparable high levels of IFN-gamma produced with 40 and 49 beads and IL-4 reactivity was still higher for larger beads (93, 101, 123 nm). Production of IgG1 was seen across the full range of bead sizes, increasing after two immunizations. Since protection against respiratory syncytial virus (RSV) depends on strong IFN responses, while IL-4 responses are reported to cause asthma-like symptoms, immunization with RSV antigens on the 49 nm carrier beads could provide the basis for a suitable vaccine. When the 49 nm beads were conjugated to RSV proteins G88 (surface) or M2.1 (internal capsid), one immunization with G88 induced high levels of IFN-gamma and low levels of IL-4. IL-4 increased with two immunizations. Beads-M2.1 induced only moderate levels of IFN-gamma and low titer antibody after two immunizations. Mice vaccinated once with G88-conjugated 49 nm beads and challenged intranasally with RSV strain A2 subtype showed reduced viral titers and recovered from weight loss more rapidly than mice immunized with M2.1-conjugated 49 nm beads or naive control mice. These results show that precise selection of nanobead size for vaccination can influence the type 1/type 2 cytokine balance after one immunization, and this will be useful in the development of effective vaccines against common human pathogens such as RSV.

Depressive symptoms in the very old living alone: prevalence, incidence and risk factors.

BACKGROUND: Living alone is one of many risk factors associated with depression. This project is nested within the ENABLE-AGE project designed to explore the relationship between housing environment and health in the very old living alone in their own homes. AIM: Our aim is to describe the prevalence, incidence and associated risk factors of clinically significant depressive symptoms in this population with particular emphasis on the role of the home environment. METHOD: We conducted a one year follow up of 376 subjects aged between 80 and 90 years old. Data collected included variables concerned with housing, social circumstances, physical health and psychological well being. RESULTS: A prevalence rate of 21% and an annual incidence of 12.4% (Geriatric Depression Score of five or more) were found. Risk factors associated with prevalence depression include not living close to friends and family ((OR 2.540, CI; 1.442, 4.466), poor satisfaction with living accommodation (OR; 0.840, CI; 0.735, 0.961) and poor satisfaction with finances (OR; 0.841, CI; 0.735, 0.961). Subsequent development of clinically significant depressive symptoms was associated with base line increased scores in depression (OR; 1.68, CI; 1.206, 2.341). CONCLUSIONS: These results are consistent with findings in the general population of similar age with the exception of considerably higher prevalence and incidence rates. However, we were unable to demonstrate that housing related variables were significant risk factors in terms of incidence cases. CLINICAL IMPLICATIONS: Older people living alone are particularly vulnerable to depression and may benefit from targeted screening and development of appropriate care pathways.

FcgammaRII and multi-system autoimmune disease.

The FcR are a crucial link in the immune response between humoral and cellular immunity and cell-based effector systems, mediating a wide variety of physiological and biochemical responses. The FcR for IgG (FcgammaR) and in particular the most widely expressed of these, FcgammaRII, are important in regulating adaptive immunity. Disruption of their function is a key factor in the development of autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), which are characterized by chronic, multi-organ inflammation. Studies of the FcgammaRII include structure/function relationships, investigation of the associations between FcR polymorphisms and human disease and animal studies using knockout or transgenic mouse models. These investigations showed that the various forms of FcgammaRII interact with immune complexes to either initiate or inhibit inflammation. In conjunction with environmental antigens and genotype, the FcgammaRII activating and inhibitory receptors determine the nature and magnitude of response to antigens. In this review, the structure and function of the FcgammaRIIs and their role in immune complex-mediated auto-immunity are discussed.

Pathogen recognition and development of particulate vaccines: does size matter?

The use of particulate carriers holds great promise for the development of effective and affordable recombinant vaccines. Rational development requires a detailed understanding of particle up-take and processing mechanisms to target cellular pathways capable of stimulating the required immune responses safely. These mechanisms are in turn based on how the host has evolved to recognize and process pathogens. Pathogens, as well as particulate vaccines, come in a wide range of sizes and biochemical compositions. Some of these also provide 'danger signals' so that antigen 'senting cells (APC), usually dendritic cells (DC), acquire specific stimulatory activity. Herein, we provide an overview of the types of particles currently under investigation for the formulation of vaccines, discuss cellular uptake mechanisms (endocytosis, macropinocytosis, phagocytosis, clathrin-dependent and/or caveloae-mediated) for pathogens and particles of different sizes, as well as antigen possessing and presentation by APC in general, and DC in particular. Since particle size and composition can influence the immune response, inducing humoral and/or cellular immunity, activating CD8 T cells and/or CD4 T cells of T helper 1 and/or T helper 2 type, particle characteristics have a major impact on vaccine efficacy. Recently developed methods for the formulation of particulate vaccines are presented in this issue of Methods, showcasing a range of "cutting edge" particulate vaccines that employ particles ranging from nano to micro-sized. This special issue of Methods further addresses practical issues of production, affordability, reproducibility and stability of formulation, and also includes a discussion of the economic and regulatory challenges encountered in developing vaccines for veterinary use and for common Third World infectious diseases.

Methods for nano-particle based vaccine formulation and evaluation of their immunogenicity.

Nano- and microparticles have long been used for the delivery of drugs and are currently being evaluated as vaccine delivery systems. Particulates can elicit potent immune responses, either by direct immuno-stimulation of antigen presenting cells (APC) or/and by delivering antigen to specific cellular compartments and promoting antigen uptake by appropriate stimulatory cell types. Herein, we describe a detailed method for the preparation of a novel nanoparticle-based antigen delivery system which induces strong cellular and humoral immune responses in mice and sheep. This simple system is based on the use of 40 nanometer (nm) inert solid carrier beads to which antigen is covalently coupled before injection. Covalent conjugation of antigen to the nanobeads, assessment of conjugation efficiency, characterisation and measurement of in vivo immunogenicity by cytokine ELISPOT (to measure antigen-specific T-cell responses) and ELISA (to measure antibody titers), are described. Emphasis is placed on providing trouble-shooting advice to enable the reproducible production of soluble nano-size formulations that do not suffer from common problems such as aggregation, as well as understanding the causes and thus avoiding a range of prevalent technical problems that occur when using immune response detection assays, such as the cytokine ELISPOT assay and ELISA.

Systemic immune responses in sheep, induced by a novel nano-bead adjuvant.

Although a number of adjuvants are currently approved for use in veterinary species, only alum has been widely used in humans. While it induces strong antibody responses, cell mediated responses are often low and inflammatory reactions at the site of injection are common. We investigated the immunological properties of a novel nano-bead adjuvant in a sheep large-animal model. In contrast to alum, antigen covalently coupled to nano-beads induced substantial cell mediated responses along with moderate humoral responses. No adverse reactions were seen at the site of immunisation in the sheep. Thus, nano-bead adjuvants in veterinary species may be useful for the induction of immunity to viral pathogens, where a cell mediated response is required. These findings also highlight the potential usefulness of nano-bead vaccines for intracellular pathogens in humans.

Development of spontaneous multisystem autoimmune disease and hypersensitivity to antibody-induced inflammation in Fcgamma receptor IIa-transgenic mice.

OBJECTIVE: The major human Fc receptor, FcgammaRIIa, is the most widespread activating FcR. Our aim was to determine the role of FcgammaRIIa in a transgenic mouse model of immune complex-mediated autoimmunity and to characterize the development of spontaneous autoimmune disease. METHODS: Arthritis was induced in normal and FcgammaRIIa-transgenic mice by immunization with type II collagen (CII) or by transfer of arthritogenic anti-CII antibodies. Also, mice that spontaneously developed autoimmune disease were assessed by clinical scoring of affected limbs, histology and serology, and measurement of autoantibody titers and cytokine production. RESULTS: FcgammaRIIa-transgenic mice developed collagen-induced arthritis (CIA) more rapidly than did archetypal CIA-sensitive DBA/1 (H-2q) mice, while nontransgenic C57BL/6 (H-2b) mice did not develop CIA when similarly immunized. Passive transfer of a single dose of anti-CII antibody induced a more rapid, severe arthritis in FcgammaRIIa-transgenic mice than in nontransgenic animals. In addition, most immune complex-induced production of tumor necrosis factor alpha by activated macrophages occurred via FcgammaRIIa, not the endogenous mouse FcR. A spontaneous, multisystem autoimmune disease developed in aging (>20 weeks) transgenic mice (n = 25), with a 32% incidence of arthritis, and by 45 weeks, all mice had developed glomerulonephritis and pneumonitis, and most had antihistone antibodies. Elevated IgG2a levels were seen in mice with CIA and in those with spontaneous disease. CONCLUSION: The presence of enhanced passive and induced autoimmunity, as well as the emergence of spontaneous autoimmune disease at 20-45 weeks of age, suggest that FcgammaRIIa is a very important factor in the pathogenesis of autoimmune inflammation and a possible target for therapeutic intervention.

Antigen delivery via two molecules on the CD8- dendritic cell subset induces humoral immunity in the absence of conventional "danger".

Targeting antigen to dendritic cells (DC) in vivo might be an effective method of modulating immune responses. Given the functional specializations among DC subsets, we investigated how targeting different receptors on different DC subsets may influence antibody (Ab) production. We show here that targeting FIRE (F4/80-like receptor) or CIRE (C-type lectin receptor), two molecules expressed on the surface of immature CD8- DC in the mouse, increases Ab production 100-1000-fold over a non-targeted control. This response was equivalent to that achieved with CpG adjuvant. In contrast, targeting CD205, which is primarily expressed on CD8+ DC, did not elicit an Ab response unless an adjuvant was added. Strong Ab responses in FcRgamma-/- mice, and with the use of F(ab')2 fragments, confirmed that FIRE and CIRE targeting was due to specific rather than FcR or complement binding. Our findings may reflect differences in the ability of CD8+ and CD8- DC subsets to stimulate immune responses in vivo. Although the consensus view is that Ag presentation on DC in their steady state leads to tolerance, the Ab enhancement from FIRE and CIRE targeting in the apparent absence of any "danger" or inflammatory signal would suggest that targeting certain DC molecules can supplant the need for external adjuvants for eliciting immune responses.

Short peptide sequences containing MHC class I and/or class II epitopes linked to nano-beads induce strong immunity and inhibition of growth of antigen-specific tumour challenge in mice.

Peptide based vaccines offer practical advantages, but unmodified peptides usually require an adjuvant or delivery vehicle to promote immunogenicity. When peptides containing ovalbumin (OVA) derived CD4 and CD8 T cell epitopes were conjugated to 0.05 microm nano-beads, they gave strong immune responses and inhibition of growth of tumour cells expressing the CD8 T cell epitope with MHC class I. These responses were inducible with both high (50 microg) and low (5 microg) peptide doses after a single immunisation. The helper CD4 T cell epitope was unnecessary for induction of CD8 T cell or tumour challenge responses. However, the CD4 T cell epitope contained a B cell epitope and triggered strong antibody responses. This simple approach offers a convenient experimental tool and a potentially useful clinical method for peptide immunisation.