Vaccine-induced neutralizing antibody responses to seasonal influenza virus H1N1 strains are not enhanced during subsequent pandemic H1N1 infection

The first exposure to influenza is presumed to shape the B-cell antibody repertoire, leading to preferential enhancement of the initially formed responses during subsequent exposure to viral variants. Here, we investigated whether this principle remains applicable when there are large genetic and antigenic differences between primary and secondary influenza virus antigens. Because humans usually have a complex history of influenza virus exposure, we conducted this investigation in influenza-naive cynomolgus macaques. Two groups of six macaques were immunized four times with influenza virus-like particles (VLPs) displaying either one (monovalent) or five (pentavalent) different hemagglutinin (HA) antigens derived from seasonal H1N1 (H1N1) strains. Four weeks after the final immunization, animals were challenged with pandemic H1N1 (H1N1pdm09). Although immunization resulted in robust virus-neutralizing responses to all VLP-based vaccine strains, there were no cross-neutralization responses to H1N1pdm09, and all animals became infected. No reductions in viral load in the nose or throat were detected in either vaccine group. After infection, strong virus-neutralizing responses to H1N1pdm09 were induced. However, there were no increases in virus-neutralizing titers against four of the five H1N1 vaccine strains; and only a mild increase was observed in virus-neutralizing titer against the influenza A/Texas/36/91 vaccine strain. After H1N1pdm09 infection, both vaccine groups showed higher virus-neutralizing titers against two H1N1 strains of intermediate antigenic distance between the H1N1 vaccine strains and H1N1pdm09, compared with the naive control group. Furthermore, both vaccine groups had higher HA-stem antibodies early after infection than the control group. In conclusion, immunization with VLPs displaying HA from antigenically distinct H1N1 variants increased the breadth of the immune response during subsequent H1N1pdm09 challenge, although this phenomenon was limited to intermediate antigenic variants

Development of broadly reactive influenza vaccines by targeting the conserved regions of the hemagglutinin stem and head domains

Introduction: Influenza virus infections cause serious illness in millions of people each year. Although influenza virus vaccines are available, they are not optimally effective due to mismatches between the influenza virus strains used for the vaccine and the circulating strains. To improve protection by vaccines, a broadly protective or universal vaccine may be required. Strategies to develop universal vaccines aim to elicit broadly reactive antibodies, which target regions on the viral hemagglutinin (HA) protein which are conserved between strains. Broadly reactive antibodies have helped to identify such targets and can guide the design of such a vaccine. Areas covered: The first part of this review provides an in-depth overview of broadly reactive anti-HA antibodies, discussing their origin, breadth and their mechanisms of protection. The second part discusses the technical design and mode of action of potential universal vaccine candidates that aim to elicit these broadly reactive antibodies and provide protection against a majority of influenza strains. Expert opinion: While great strides have been made in the development of universal influenza vaccine candidates, real-life use still requires improvement of stability, enhancement of their breadth of protection and ease of production, while efficacies need to be determined in human trials

Quantitative in vitro-to-in vivo extrapolation (QIVIVE) of estrogenic and anti-androgenic potencies of BPA and BADGE analogues

The goal of the present study was to obtain an in vivo relevant prioritization method for the endocrine potencies of different polycarbonate monomers, by combining in vitro bioassay data with physiologically based kinetic (PBK) modelling. PBK models were developed for a selection of monomers, including bisphenol A (BPA), two bisphenol F (BPF) isomers and four different bisphenol A diglycidyl ethers (BADGEs), using in vitro input data. With these models, the plasma concentrations of the compounds were simulated, providing means to estimate the dose levels at which the in vitro endocrine effect concentrations are reached. The results revealed that, whereas the in vitro relative potencies of different BADGEs (predominantly anti-androgenic effects) can be up to fourfold higher than BPA, the estimated in vivo potencies based on the oral equivalent doses are one to two orders of magnitude lower than BPA because of fast detoxification of the BADGEs. In contrast, the relative potencies of 2,2-BPF and 4,4-BPF increase when accounting for the in vivo availability. 4,4-BPF is estimated to be fivefold more potent than BPA in humans in vivo in inducing estrogenic effects and both 2,2-BPF and 4,4-BPF are estimated to be, respectively, 7 and 11-fold more potent in inducing anti-androgenic effects. These relative potencies were considered to be first-tier estimates, particularly given that the potential influence of intestinal metabolism on the in vivo availability was not accounted for. Overall, it can be concluded that both 2,2-BPF and 4,4-BPF are priority compounds

Influenza a virus hemagglutinin trimer, head and stem proteins identify and quantify different hemagglutinin-specific b cell subsets in humans

Antibody responses against the influenza A virus hemagglutinin (HA)-protein are studied intensively because they can protect against (re)infection. Previous studies have focused on antibodies targeting the head or stem domains, while other possible specificities are often not taken into account. To study such specificities, we developed a diverse set of HA-domain proteins based on an H1N1pdm2009-like influenza virus strain, including monomeric head and trimeric stem domain, as well as the full HA-trimer. These proteins were used to study the B cell and antibody responses in six healthy human donors. A large proportion of HA-trimer B cells bound exclusively to HA-trimer probe (54–77%), while only 8–18% and 9–23% were able to recognize the stem or head probe, respectively. Monoclonal antibodies (mAbs) were isolated and three of these mAbs, targeting the different domains, were characterized in-depth to confirm the binding profile observed in flow cytometry. The head-directed mAb, targeting an epitope distinct from known head-specific mAbs, showed relatively broad H1N1 neutralization and the stem-directed mAb was able to broadly neutralize diverse H1N1 viruses. Moreover, we identified a trimer-directed mAb that did not compete with known head or stem domain specific mAbs, suggesting that it targets an unknown epitope or conformation of influenza virus’ HA. These observations indicate that the described method can characterize the diverse antibody response to HA and might be able to identify HA-specific B cells and antibodies with previously unknown specificities that could be relevant for vaccine design

Evaluation of a loop-mediated isothermal amplification (LAMP) method for rapid on-site detection of horse meat

Detection of horse DNA by loop-mediated isothermal amplification (LAMP) seems one of the most promising methods to meet the criteria of fast, robust, cost efficient, specific, and sensitive on-site detection. In the present study an assessment of the specificity and sensitivity of the LAMP horse screening assay was made and outcomes were compared with the EURL-AP (European Union Reference laboratory for Animal Proteins in feeding stuffs) qPCR method. The specificity was tested with DNA samples from seven other species. The sensitivity of the LAMP assay was subsequently challenged with different percentages of horse DNA in cattle DNA and different percentages of horse meat in cattle meat. Both qPCR and LAMP were able to reliably detect horse DNA or meat below 0.1%, but LAMP was able to do so in less than 30 min. The DNA of other species did not result in a response in the LAMP horse assay. These features show that the LAMP method is fast, specific, and sensitive. Next, 69 processed meat samples were screened for the presence of horse DNA. The results showed that the LAMP horse assay, combined with a simple and fast on-site DNA extraction method, results in similar outcomes as the EURL-AP qPCR method and is thus a promising screening assay to be used outside the laboratory. Only samples that are screened on-site as suspect in the LAMP horse assay, need to be brought to the laboratory for confirmation with the more laborious EURL-AP qPCR reference method.</p

Evaluation of a loop-mediated isothermal amplification (LAMP) method for rapid on-site detection of horse meat

Detection of horse DNA by loop-mediated isothermal amplification (LAMP) seems one of the most promising methods to meet the criteria of fast, robust, cost efficient, specific, and sensitive on-site detection. In the present study an assessment of the specificity and sensitivity of the LAMP horse screening assay was made and outcomes were compared with the EURL-AP (European Union Reference laboratory for Animal Proteins in feeding stuffs) qPCR method. The specificity was tested with DNA samples from seven other species. The sensitivity of the LAMP assay was subsequently challenged with different percentages of horse DNA in cattle DNA and different percentages of horse meat in cattle meat. Both qPCR and LAMP were able to reliably detect horse DNA or meat below 0.1%, but LAMP was able to do so in less than 30 min. The DNA of other species did not result in a response in the LAMP horse assay. These features show that the LAMP method is fast, specific, and sensitive. Next, 69 processed meat samples were screened for the presence of horse DNA. The results showed that the LAMP horse assay, combined with a simple and fast on-site DNA extraction method, results in similar outcomes as the EURL-AP qPCR method and is thus a promising screening assay to be used outside the laboratory. Only samples that are screened on-site as suspect in the LAMP horse assay, need to be brought to the laboratory for confirmation with the more laborious EURL-AP qPCR reference method.</p

Boletín de Segovia: Número 3 - 1913 enero 6

Copia digital. Madrid : Ministerio de Cultura. Subdirección General de Coordinación Bibliotecaria, 200

Additional file 8: Figure S6. of Glucocorticoid receptor and nuclear factor kappa-b affect three-dimensional chromatin organization

(A) Histogram depicting the genomic proximity (localization) of P300 binding sites identified by ChIP-seq in relation to those identified by using ChIA-PET self-ligation PETs. Identical comparison is performed for both DMSO-treated and TA + TNFα-treated data sets. (B) P300 ChIP-seq signal at P300 binding sites commonly identified by ChIP-seq and ChIA-PET and those binding sites that were uniquely detected in the ChIP-seq data set. (C) P300 ChIP-seq signal at P300 binding sites that were either involved (anchor) or not involved (non-anchor) in long-range interaction as identified by ChIA-PET analysis. (D) An example screenshot depicting the P300 interaction subdomains, P300 ChIP-seq binding sites in relation to topological domains as defined by replication timing data ( www.replicationdomain.org ). (E) Localization of all the interaction subdomains identified by ChIA-PET analysis (P300 and POLII) in relation to topological domains. (PDF 1041 kb

Additional file 11: Figure S8. of Glucocorticoid receptor and nuclear factor kappa-b affect three-dimensional chromatin organization

Direct comparison of long-range interactions identified by P300 ChIA-PET and 4C-seq analyses at FKBP5, DUSP1, and BIRC3 loci. (PDF 1037 kb

Additional file 2: Figure S2. of Glucocorticoid receptor and nuclear factor kappa-b affect three-dimensional chromatin organization

Activated p65 induces de novo P300 depositions to latent genomic loci. (A) Pile-up heat map depicting the H3K4me1 and H3K4me3 signal around (±12 kb) all p65-bound promoters and DBSs. (B) Pile-up heat map depicting the p65 and P300 signal at all p65-bound enhancers upon vehicle, DMSO (−), and TNFα (+) treatment. (C) Example screenshot depicting TNFα-induced P300 recruitment at genomic regions (red box) and recruitment of p65 at genomic loci that are pre-marked by P300. (D) Motif occurrence at all p65-bound DBS presented as a function of TNFα-dependent P300 recruitment (x-axis) (top-panel). Level of shared binding of p65 and other TFs at all p65-bound DBS, presented as a function of TNFα-dependent P300 recruitment (bottom panel). (E) Level of H3K27ac, DNase I hypersensitivity, and H3K4me1 at all p65-bound DBSs (induced and constitutive P300 sites). (PDF 909 kb