Systemic and Mucosal Immune Responses to Sublingual or Intramuscular Human Papilloma Virus Antigens in Healthy Female Volunteers

The sublingual route has been proposed as a needle-free option to induce systemic and mucosal immune protection against viral infections. In a translational study of systemic and mucosal humoral immune responses to sublingual or systemically administered viral antigens, eighteen healthy female volunteers aged 19–31 years received three immunizations with a quadravalent Human Papilloma Virus vaccine at 0, 4 and 16 weeks as sublingual drops (SL, n = 12) or intramuscular injection (IM, n = 6). IM antigen delivery induced or boosted HPV-specific serum IgG and pseudovirus-neutralizing antibodies, HPV-specific cervical and vaginal IgG, and elicited circulating IgG and IgA antibody secreting cells. SL antigens induced ∼38-fold lower serum and ∼2-fold lower cervical/vaginal IgG than IM delivery, and induced or boosted serum virus neutralizing antibody in only 3/12 subjects. Neither route reproducibly induced HPV-specific mucosal IgA. Alternative delivery systems and adjuvants will be required to enhance and evaluate immune responses following sublingual immunization in humans

Changes in cytokine production in healthy subjects practicing Guolin Qigong : a pilot study

BACKGROUND: Guolin Qigong is a combination of meditation, controlled breathing and physical movement designed to control the vital energy (qi) of the body and consequently to improve spiritual, physical and mental health. Practice of Qigong has been reported to alter immunological function, but there have been few studies of its effects on cytokines, the key regulators of immunity. METHODS: Numbers of peripheral blood cytokine-secreting cells were determined by ELISPOT in 19 healthy volunteers aged 27 – 55, before they were taught the practice of Qigong and after 3, 7 and 14 weeks of daily practice. The effect of Qigong on blood cortisol was also examined. RESULTS: Numbers of IL4 and IL12-secreting cells remained stable. IL6 increased at 7 weeks and TNFα increased in unstimulated cultures at 3 and 7 weeks but decreased at these times in LPS and SAC-stimulated cultures. Of particular interest, IFNγ-secreting cells increased and IL10-secreting cells decreased in PHA-stimulated cultures, resulting in significant increases in the IFNγ:IL10 ratio. Cortisol, a known inhibitor of type 1 cytokine production, was reduced by practicing Qigong. CONCLUSION: These preliminary studies in healthy subjects, although not necessarily representative of a randomized healthy population and not including a separate control group, have indicated that blood levels of the stress-related hormone cortisol may be lowered by short-term practice of Qigong and that there are concomitant changes in numbers of cytokine-secreting cells. Further studies of the effect of Qigong in patients with clinical diseases known to be associated with type 2 cytokine predominance are merited

Galectin-8 in IgA Nephritis: Decreased Binding of IgA by Galectin-8 Affinity Chromatography and Associated Increased Binding in Non-IgA Serum Glycoproteins

Background Immunoglobulin A nephritis (IgAN) is the most common primary glomerulonephritis worldwide. It is caused by accumulation of IgA1-containing immune complexes in the kidney resulting in renal failure, which is thought to be due to altered glycosylation of IgA with a decrease of 2-3-sialylated galactosides (NeuAc alpha 2-3Gal). less thanbrgreater than less thanbrgreater thanPurpose The purpose of this study was to analyze whether altered glycosylation of IgA would lead to an altered binding to galectin-8, an endogenous lectin with strong affinity for 2-3-sialylated galactosides. Galectins are a family of beta-galactoside-binding proteins; by binding various glycoproteins, they play important roles in the regulation of cellular functions in inflammation and immunity. Hence, an altered binding of IgA to galectin-8 could lead to pathologic immune functions, such as glomerulonephritis. less thanbrgreater than less thanbrgreater thanMethods Affinity chromatography of serum glycoproteins on the human sialogalactoside-binding lectin galectin-8N permitted quantitation of bound and unbound fractions, including IgA. less thanbrgreater than less thanbrgreater thanResults Analysis of similar to 100 IgA nephritis sera showed that the galectin-8N unbound fraction of IgA increased compared to similar to 100 controls, consistent with the known loss of galactosylation. A subgroup of similar to 15% of the IgAN patients had a ratio of galectin-8 bound/unbound IgA andlt;0.09, not found for any of the controls. Unexpectedly, the galectin-8N-binding fraction of serum glycoproteins other than IgA increased in the sera of IgAN patients but not in controls, suggesting a previously unrecognized change in this disease. less thanbrgreater than less thanbrgreater thanConclusion This is the first study that relates a galectin, an endogenous lectin family, to IgA nephritis and thus should stimulate new avenues of research into the pathophysiology of the disease.Funding Agencies|Swedish Research Council (Vetenskapsradet)|2008-3356|Swedish Foundation for Swedish Research|FFL4|Swedish Healthcare System (ALF)||Region Skane||</p

Peptide microarrays for the profiling of cytotoxic T-lymphocyte activity using minimum numbers of cells

The identification of epitopes that elicit cytotoxic T-lymphocyte activity is a prerequisite for the development of cancer-specific immunotherapies. However, especially the parallel characterization of several epitopes is limited by the availability of T cells. Microarrays have enabled an unprecedented miniaturization and parallelization in biological assays. Here, we developed peptide microarrays for the detection of CTL activity. MHC class I-binding peptide epitopes were pipetted onto polymer-coated glass slides. Target cells, loaded with the cell-impermeant dye calcein, were incubated on these arrays, followed by incubation with antigen-expanded CTLs. Cytotoxic activity was detected by release of calcein and detachment of target cells. With only 200,000 cells per microarray, CTLs could be detected at a frequency of 0.5% corresponding to 1,000 antigen-specific T cells. Target cells and CTLs only settled on peptide spots enabling a clear separation of individual epitopes. Even though no physical boundaries were present between the individual spots, peptide loading only occurred locally and cytolytic activity was confined to the spots carrying the specific epitope. The peptide microarrays provide a robust platform that implements the whole process from antigen presentation to the detection of CTL activity in a miniaturized format. The method surpasses all established methods in the minimum numbers of cells required. With antigen uptake occurring on the microarray, further applications are foreseen in the testing of antigen precursors that require uptake and processing prior to presentation

In vitro and in vivo mRNA delivery using lipid-enveloped pHresponsive polymer nanoparticles

Biodegradable core−shell structured nanoparticles with a poly(β-amino ester) (PBAE) core enveloped by a phospholipid bilayer shell were developed for in vivo mRNA delivery with a view toward delivery of mRNA-based vaccines. The pH-responsive PBAE component was chosen to promote endosome disruption, while the lipid surface layer was selected to minimize toxicity of the polycation core. Messenger RNA was efficiently adsorbed via electrostatic interactions onto the surface of these net positively charged nanoparticles. In vitro, mRNA-loaded particle uptake by dendritic cells led to mRNA delivery into the cytosol with low cytotoxicity, followed by translation of the encoded protein in these difficult-to-transfect cells at a frequency of 30%. Particles loaded with mRNA administered intranasally (i.n.) in mice led to the expression of the reporter protein luciferase in vivo as soon as 6 h after administration, a time point when naked mRNA given i.n. showed no expression. At later time points, luciferase expression was detected in naked mRNA-treated mice, but this group showed a wide variation in levels of transfection, compared to particle-treated mice. This system may thus be promising for noninvasive delivery of mRNA-based vaccines.United States. Dept. of Defense (Institute for Soldier Nanotechnology, contract W911NF-07-D-0004)Ragon Institute of MGH, MIT and HarvardSingapore. Agency for Science, Technology and ResearchHoward Hughes Medical Institute (Investigator

Oral Immunization with a Live Coxsackievirus/HIV Recombinant Induces Gag p24-Specific T Cell Responses

The development of an HIV/AIDS vaccine has proven to be elusive. Because human vaccine trials have not yet demonstrated efficacy, new vaccine strategies are needed for the HIV vaccine pipeline. We have been developing a new HIV vaccine platform using a live enterovirus, coxsackievirus B4 (CVB4) vector. Enteroviruses are ideal candidates for development as a vaccine vector for oral delivery, because these viruses normally enter the body via the oral route and survive the acidic environment of the stomach.We constructed a live coxsackievirus B4 recombinant, CVB4/p24(73(3)), that expresses seventy-three amino acids of the gag p24 sequence (HXB2) and assessed T cell responses after immunization of mice. The CVB4 recombinant was physically stable, replication-competent, and genetically stable. Oral or intraperitoneal immunization with the recombinant resulted in strong systemic gag p24-specific T cell responses as determined by the IFN-gamma ELISPOT assay and by multiparameter flow cytometry. Oral immunization with CVB4/p24(73(3)) resulted in a short-lived, localized infection of the gut without systemic spread. Because coxsackieviruses are ubiquitous in the human population, we also evaluated whether the recombinant was able to induce gag p24-specific T cell responses in mice pre-immunized with the CVB4 vector. We showed that oral immunization with CVB4/p24(73(3)) induced gag p24-specific immune responses in vector-immune mice.The CVB4/p24(73(3)) recombinant retained the physical and biological characteristics of the parental CVB4 vector. Oral immunization with the CVB4 recombinant was safe and resulted in the induction of systemic HIV-specific T cell responses. Furthermore, pre-existing vector immunity did not preclude the development of gag p24-specific T cell responses. As the search continues for new vaccine strategies, the present study suggests that live CVB4/HIV recombinants are potential new vaccine candidates for HIV

Immunogenicity and efficacy of oral vaccines in developing countries: lessons from a live cholera vaccine

Oral vaccines, whether living or non-living, viral or bacterial, elicit diminished immune responses or have lower efficacy in developing countries than in developed countries. Here I describe studies with a live oral cholera vaccine that include older children no longer deriving immune support from breast milk or maternal antibodies and that identify some of the factors accounting for the lower immunogenicity, as well as suggesting counter-measures that may enhance the effectiveness of oral immunization in developing countries. The fundamental breakthrough is likely to require reversing effects of the 'environmental enteropathy' that is often present in children living in fecally contaminated, impoverished environments