A Recombinant Vaccine Effectively Induces C5a-Specific Neutralizing Antibodies and Prevents Arthritis

OBJECTIVES: To develop and validate a recombinant vaccine to attenuate inflammation in arthritis by sustained neutralization of the anaphylatoxin C5a. METHODS: We constructed and expressed fusion protein of C5a and maltose binding protein. Efficacy of specific C5a neutralization was tested using the fusion protein as vaccine in three different arthritis mouse models: collagen induced arthritis (CIA), chronic relapsing CIA and collagen antibody induced arthritis (CAIA). Levels of anti-C5a antibodies and anti-collagen type II were measured by ELISA. C5a neutralization assay was done using a rat basophilic leukemia cell-line transfected with the human C5aR. Complement activity was determined using a hemolytic assay and joint morphology was assessed by histology. RESULTS: Vaccination of mice with MBP-C5a led to significant reduction of arthritis incidence and severity but not anti-collagen antibody synthesis. Histology of the MBP-C5a and control (MBP or PBS) vaccinated mice paws confirmed the vaccination effect. Sera from the vaccinated mice developed C5a-specific neutralizing antibodies, however C5 activation and formation of the membrane attack complex by C5b were not significantly altered. CONCLUSIONS: Exploitation of host immune response to generate sustained C5a neutralizing antibodies without significantly compromising C5/C5b activity is a useful strategy for developing an effective vaccine for antibody mediated and C5a dependent inflammatory diseases. Further developing of such a therapeutic vaccine would be more optimal and cost effective to attenuate inflammation without affecting host immunity

Development of a New Oral Vaccine against Diphtheria and the Study of its Immunogenicity in Mouse and Man

Most pathogens enter the body via mucosal surfaces. In contrast to parenterally administered vaccination, mucosal vaccination has the advantage of eliciting both a systemic and a local mucosal immune response. An oral biodegradable adjuvant with these features would have great potential. This thesis has focused on the development of a new oral vaccine against diphtheria. Biodegradable polyacryl starch microparticles were used as a mucosal adjuvant. Diphtheria toxin or cross-reacting material of diphtheria toxin (CRM197) was covalently conjugated to the microparticles and fed to mice by oral gavage. Formaldehyde treatment was also studied as a means of either detoxifying (diphtheria toxin) or stabilising (CRM197) these formulations. All formulations given to mice orally or parenterally, but not intranasally, induced a strong systemic immune response and diphtheria toxin neutralising antibodies. Only formulations administered orally induced a mucosal IgA response as well. The non-toxic recombinant protein CRM197 proved to be a promising antigen candidate in an oral diphtheria vaccine when conjugated to the microparticles. Mild treatment of CRM197 with formaldehyde before conjugation to the starch microparticles potentiated the immunogenicity of the formulation. However, no immune response was detected in healthy volunteers after administration of this vaccine in a phase I trial. The possible reasons for the difference in response between mouse and man are discussed. The use of cDNA expression macro array technology was also evaluated as a tool in vaccine-related research. Tetanus toxoid and aluminium phosphate were used as model parenteral antigen and adjuvant. It was concluded that the antigen modulates the molecular mechanisms of the aluminium phosphate adjuvant to a greater extent than previously recognised

Measurement of Serum IgG Anti-Integrin alpha v beta 6 Autoantibodies Is a Promising Tool in the Diagnosis of Ulcerative Colitis

IgG anti-integrin alpha v beta 6 autoantibodies (IgG anti-alpha v beta 6) have been described as highly sensitive and specific markers of ulcerative colitis (UC) in the sera of Japanese inflammatory bowel disease (IBD) patients. We aimed to evaluate the diagnostic performance of IgG anti-alpha v beta 6 as a biomarker in Swedish patients with IBD or irritable bowel syndrome (IBS). The study included adult UC (n = 59), Crohn's disease (CD, n = 38), and IBS patients (n = 100). Partial Mayo score and Harvey-Bradshaw index were used to assess disease severity for UC and CD, respectively. Serum levels of IgG anti-alpha v beta 6, reported as absorbance units (AU), were measured using an in-house ELISA where the 95th percentile of 76 healthy controls defined positivity. Faecal calprotectin (fCP) was measured using a commercial assay. The majority of the IBD patients were on medical treatment, and many were in remission (UC: 40.7%; CD: 47.4%). Seventy-one percent of the UC patients, 74.2% of CD patients, and 23.1% of the IBS patients had fCP test results >50 mg/kg. The UC group had significantly higher IgG anti-alpha v beta 6 levels (median: 1.76 AU) than the CD and IBS groups (0.34 and 0.31 AU, both p < 0.0001). The diagnostic sensitivity of IgG anti-alpha v beta 6 in UC was 76.3%, and the specificities were 79.0% (vs. CD) and 96.0% (vs. IBS). The IgG anti-alpha v beta 6 levels related to disease severity of the UC patients (p < 0.01-0.05). Our study shows that IgG anti-alpha v beta 6 is associated with UC in Swedish IBD patients and that the levels of the autoantibodies reflect disease severity. IgG anti-alpha v beta 6 could be an attractive complement to fCP in the diagnostic work up of IBD patients

A comparison of visual ergonomic measurements between active and passive 3DTV

The two latest 3DTV types were tested; active shutter eye-glass type and passive eye-glass type with screen having film pattern retarder. Differences in vertical resolution, cross-talk and luminance were found. This difference in vertical resolution is visible for some types of objects. How this affects the image quality is unclear

Clinical potential of eosinophil-derived neurotoxin in asthma management

The assessment and management of asthma patients is challenging because of the complexity of the underlying inflammatory mechanisms and heterogeneity of their clinical presentation. Optimizing disease management requires therapy individualization that should rely on reliable biomarkers to unravel the phenotypes and endotypes of asthma. The secretory activity and turnover of eosinophils, as assessed by measuring eosinophil-derived proteins, may provide an accurate and complementary tool that mirrors the eosinophil activation status. Emerging evidence suggests that eosinophil-derived neurotoxin (EDN) has considerable potential as a precision medicine biomarker. In this review, we explore EDN suitability as biomarker in asthma management, with particular emphasis on its clinical significance in the management of both pediatric and adult populations

Tryptase reference values in a Swedish middle-aged general population and association with diabetes mellitus

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Histology of joint sections.

<p>Effect of C5a inhibition in CIA (A–C) and CAIA (D–E) demonstrated in representative tissue histology. 4–5 mice in each group from the experiments shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013511#pone-0013511-g001" target="_blank">Fig. 1B</a> (CIA) and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013511#pone-0013511-g003" target="_blank">Fig. 3</a> (CAIA) were sacrificed and joint morphology was assessed using a standard haematoxylin/eosin staining protocol. Representative joint sections (6 µm) from QB mice induced with CIA but vaccinated with PBS (A), MBP (B) or MBP-C5a (C) are shown. Magnification x10. Joint sections from mice induced with CAIA and vaccinated with MBP-C5a (D) or control (PBS) (E) are shown. Magnification x20.</p

Measurement of C5 activity in mouse sera.

<p>Sera were collected on indicated days from mice described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013511#pone-0013511-g001" target="_blank">Fig. 1B</a> (CIA), <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013511#pone-0013511-g003" target="_blank">Fig. 3</a> (CAIA) and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013511#pone-0013511-g004" target="_blank">Fig. 4D</a> (C5 sufficient B10.Q and C5 deficient NB2 mice). Serum samples were analyzed for C5 activity A) on days −21 and 35 from MBP-C5a, MBP or PBS vaccinated mice used in CIA experiment, B) day 0 and 35 from MBP-C5a or PBS vaccinated mice used in CAIA experiment and C) day −21, 34 and 90 from MBP-C5a or PBS vaccinated C5 sufficient B10.Q or C5 deficient NB2 congenic mice. Great care was taken in order to preserve complement activity. Activity of the alternative pathway of complement was measured in hemolytic assay in which rabbit erythrocytes were incubated with mouse sera and the amount of hemoglobin released in supernatants due to lysis was determined spectrophotometrically. All the assays were done in triplicates. There was no significant difference in complement activity between MBP-C5a and MBP control (A) but there was a significant difference in complement activity when MBP-C5a and PBS groups were compared (in Fig. A and B but not in C) using t-test (paired). Error bars indicate ± SD.</p