Transplacentally Acquired Maternal Antibody against Hepatitis B Surface Antigen in Infants and its Influence on the Response to Hepatitis B Vaccine

BACKGROUND: Passively acquired maternal antibodies in infants may inhibit active immune responses to vaccines. Whether maternal antibody against hepatitis B surface antigen (anti-HBs) in infants may influence the long-term immunogenicity of hepatitis B vaccine remains unknown. METHODOLOGY/PRINCIPAL FINDINGS: Totally 338 pairs of mothers and children were enrolled. All infants were routinely vaccinated against hepatitis B based on 0-, 1- and 6-month schedule. We characterized the transplacental transfer of maternal anti-HBs, and compared anti-HBs response in children of mothers with or without anti-HBs. In a prospective observation, all 63 anti-HBs positive mothers transferred anti-HBs to their infants; 84.1% of the infants had higher anti-HBs concentrations than their mothers. One and half years after vaccination with three doses of hepatitis B vaccine, the positive rate and geometric mean concentration (GMC) of anti-HBs in 32 infants with maternal anti-HBs were comparable with those in 32 infants without maternal antibody (90.6% vs 87.5%, P = 0.688, and 74.5 vs 73.5 mIU/ml, P = 0.742, respectively). In a retrospective analysis, five and half years after vaccination with three doses vaccine, the positive rates of anti-HBs in 88 children of mothers with anti-HBs ≥1000 mIU/ml, 94 children of mothers with anti-HBs 10-999 mIU/ml, and 61 children of mothers with anti-HBs <10 mIU/ml were 72.7%, 69.2%, and 63.9% (P = 0.521), respectively; anti-HBs GMC in these three groups were 38.9, 43.9, and 31.7 mIU/ml (P = 0.726), respectively. CONCLUSIONS/SIGNIFICANCE: The data demonstrate that maternal anti-HBs in infants, even at high concentrations, does not inhibit the long-term immunogenicity of hepatitis B vaccine. Thus, current hepatitis B vaccination schedule for infants will be still effective in the future when most infants are positive for maternal anti-HBs due to the massive vaccination against hepatitis B

FcRn-mediated antibody transport across epithelial cells revealed by electron tomography

The neonatal Fc receptor (FcRn) transports maternal IgG across epithelial barriers, thereby providing the fetus or newborn with humoral immunity before its immune system is fully functional. In newborn rats, FcRn transfers IgG from milk to blood by apical-to-basolateral transcytosis across intestinal epithelial cells. The pH difference between the apical (pH 6.0–6.5) and basolateral (pH 7.4) sides of intestinal epithelial cells facilitates the efficient unidirectional transport of IgG, because FcRn binds IgG at pH 6.0–6.5 but not at pH 7 or more. As milk passes through the neonatal intestine, maternal IgG is removed by FcRn-expressing cells in the proximal small intestine (duodenum and jejunum); remaining proteins are absorbed and degraded by FcRn-negative cells in the distal small intestine (ileum). Here we use electron tomography to make jejunal transcytosis visible directly in space and time, developing new labelling and detection methods to map individual nanogold-labelled Fc within transport vesicles and simultaneously to characterize these vesicles by immunolabelling. Combining electron tomography with a nonperturbing endocytic label allowed us to conclusively identify receptor-bound ligands, resolve interconnecting vesicles, determine whether a vesicle was microtubule-associated, and accurately trace FcRn-mediated transport of IgG. Our results present a complex picture in which Fc moves through networks of entangled tubular and irregular vesicles, only some of which are microtubule-associated, as it migrates to the basolateral surface. New features of transcytosis are elucidated, including transport involving multivesicular body inner vesicles/tubules and exocytosis through clathrin-coated pits. Markers for early, late and recycling endosomes each labelled vesicles in different and overlapping morphological classes, revealing spatial complexity in endo-lysosomal trafficking

Characterization of the Rabbit Neonatal Fc Receptor (FcRn) and Analyzing the Immunophenotype of the Transgenic Rabbits That Overexpresses FcRn

The neonatal Fc receptor (FcRn) regulates IgG and albumin homeostasis, mediates maternal IgG transport, takes an active role in phagocytosis, and delivers antigen for presentation. We have previously shown that overexpression of FcRn in transgenic mice significantly improves the humoral immune response. Because rabbits are an important source of polyclonal and monoclonal antibodies, adaptation of our FcRn overexpression technology in this species would bring significant advantages. We cloned the full length cDNA of the rabbit FcRn alpha-chain and found that it is similar to its orthologous analyzed so far. The rabbit FcRn - IgG contact residues are highly conserved, and based on this we predicted pH dependent interaction, which we confirmed by analyzing the pH dependent binding of FcRn to rabbit IgG using yolk sac lysates of rabbit fetuses by Western blot. Using immunohistochemistry, we detected strong FcRn staining in the endodermal cells of the rabbit yolk sac membrane, while the placental trophoblast cells and amnion showed no FcRn staining. Then, using BAC transgenesis we generated transgenic rabbits carrying and overexpressing a 110 kb rabbit genomic fragment encoding the FcRn. These transgenic rabbits – having one extra copy of the FcRn when hemizygous and two extra copies when homozygous - showed improved IgG protection and an augmented humoral immune response when immunized with a variety of different antigens. Our results in these transgenic rabbits demonstrate an increased immune response, similar to what we described in mice, indicating that FcRn overexpression brings significant advantages for the production of polyclonal and monoclonal antibodies

FcRn Overexpression in Transgenic Mice Results in Augmented APC Activity and Robust Immune Response with Increased Diversity of Induced Antibodies

Our previous studies have shown that overexpression of bovine FcRn (bFcRn) in transgenic (Tg) mice leads to an increase in the humoral immune response, characterized by larger numbers of Ag-specific B cells and other immune cells in secondary lymphoid organs and higher levels of circulating Ag-specific antibodies (Abs). To gain additional insights into the mechanisms underlying this increase in humoral immune response, we further characterized the bFcRn Tg mice. Our Western blot analysis showed strong expression of the bFcRn transgene in peritoneal macrophages and bone marrow derived dendritic cells; and a quantitative PCR analysis demonstrated that the expression ratios of the bFcRn to mFcRn were 2.6- and 10-fold in these cells, respectively. We also found that overexpression of bFcRn enhances the phagocytosis of Ag-IgG immune complexes (ICs) by both macrophages and dendritic cells and significantly improves Ag presentation by dendritic cells. Finally, we determined that immunized bFcRn mice produce a much greater diversity of Ag-specific IgM, whereas only the levels, but not the diversity, of IgG is increased by overexpression of bFcRn. We suggest that the increase in diversity of IgG in Tg mice is prevented by a selective bias towards immunodominant epitopes of ovalbumin, which was used in this study as a model antigen. These results are also in line with our previous reports describing a substantial increase in the levels of Ag-specific IgG in FcRn Tg mice immunized with Ags that are weakly immunogenic and, therefore, not affected by immunodominance

Spotlight on Differentially Expressed Genes in Urinary Bladder Cancer

INTRODUCTION: We previously identified common differentially expressed (DE) genes in bladder cancer (BC). In the present study we analyzed in depth, the expression of several groups of these DE genes. MATERIALS AND METHODS: Samples from 30 human BCs and their adjacent normal tissues were analyzed by whole genome cDNA microarrays, qRT-PCR and Western blotting. Our attention was focused on cell-cycle control and DNA damage repair genes, genes related to apoptosis, signal transduction, angiogenesis, as well as cellular proliferation, invasion and metastasis. Four publicly available GEO Datasets were further analyzed, and the expression data of the genes of interest (GOIs) were compared to those of the present study. The relationship among the GOI was also investigated. GO and KEGG molecular pathway analysis was performed to identify possible enrichment of genes with specific biological themes. RESULTS: Unsupervised cluster analysis of DNA microarray data revealed a clear distinction in BC vs. control samples and low vs. high grade tumors. Genes with at least 2-fold differential expression in BC vs. controls, as well as in non-muscle invasive vs. muscle invasive tumors and in low vs. high grade tumors, were identified and ranked. Specific attention was paid to the changes in osteopontin (OPN, SPP1) expression, due to its multiple biological functions. Similarly, genes exhibiting equal or low expression in BC vs. the controls were scored. Significant pair-wise correlations in gene expression were scored. GO analysis revealed the multi-facet character of the GOIs, since they participate in a variety of mechanisms, including cell proliferation, cell death, metabolism, cell shape, and cytoskeletal re-organization. KEGG analysis revealed that the most significant pathway was that of Bladder Cancer (p = 1.5×10(-31)). CONCLUSIONS: The present work adds to the current knowledge on molecular signature identification of BC. Such works should progress in order to gain more insight into disease molecular mechanisms

Interaction between gelatin and sodium dodecyl sulfate at the air/water interface: A neutron reflection study

Neutron reflection has been used to study the composition and structure of layers adsorbed at the air/water surface of solutions of gelatin and sodium dodecyl sulfate (SDS) and these results have been compared with the surface tension of the same solutions. Above a concentration where free micelles of SDS can be expected to form in the bulk solution the layer is exactly as would be expected for solutions of SDS on its own. However, at low SDS concentrations the presence of gelatin greatly enhances the adsorption of SDS in comparison with solutions just containing surfactant, and in the intermediate range of SDS concentration, where the surface tension is relatively constant, the surface excess of SDS is also constant at gelatin concentrations of 0.1 wt%. The thickness of the surfactant layer in the two lower ranges of SDS concentration is much larger than a simple surfactant layer, ranging from 35 down to 22 angstroms (in comparison with 19 angstroms for the pure surfactant layer), suggesting that the layer is not only roughened by binding of gelatin at the surface but that a proportion of the bound SDS molecules are completely immersed just below the surface. This is confirmed by measurements of the layer structure at different isotopic compositions. The presence of gelatin at the surface and the enhancement of the adsorption of SDS indicate that complexes of gelatin and SDS are strongly surface active. Furthermore, the measured thickness of the SDS layer at the surface shows that these complexes probably do not contain surfactant in the form of micelles. This further suggests that it may not be reliable to interpret the first discontinuity in the surface tensionlog(concentration) plot in such strongly interacting systems as the point at which there is an onset in aggregation of the surfactant on the polyelectrolyte (critical aggregation concentration, or CAC)

Neutron reflection study of a double-Chained sugar surfactant

Neutron reflection and surface tension have been used to investigate the structural features exhibited at the air-liquid interface by an adsorbed layer of the sugar-based nonionic surfactant 7,7-bis[(1,2,3,4,5-pentahydroxyhexanamido)methyl]-n-tridecane. It consists of two glucamide headgroups and two C16H13 alkyl chains, i.e., (C6H13)2C(CH2NHCO(CHOH) 4CH2OH)2, abbreviated to di(C6-Glu). Isotopic substitution was used to distinguish the chains of the surfactant from the headgroup and solvent distributions. The critical micelle concentration (CMC) was found to be (1.9 ± 0.1) × 10-3 M using surface tension measurements, and the latter also indicated that the surface properties were not affected by isotopic substitution. The specular reflection of neutrons has been used to determine the surface concentrations of skeletally deuterated di(C6-Glu) over the concentration range CMC/400 to 2CMC. The neutron measurements showed the area per molecule A at the CMC to be 66 ± 3 Å2, as compared with 60 ± 10 Å2 from surface tension measurements. The layer structure has been determined using three different isotopic contrasts at concentrations of CMC and CMC/400. The fit of a two-layer model shows that the total monolayer thickness decreases from 17.5 ± 1.5 Å at the CMC to 14.5 ± 1.5 Å at CMC/400, respectively. The kinematic approximation was used to fit the neutron data and an X-ray reflection profile. The most remarkable feature of the structure obtained is the apparent smoothness of the layer. The calculated roughness of the layer at the CMC is 9.7 Å2 which corresponds exactly to the value expected for capillary waves using the model of a pure liquid. At the CMC the molecule as a whole appears to be tilted about 35-40° away from the surface normal, but at CMC/400 the hydrocarbon chains are approximately normal to the surface while the gluconamide groups are strongly tilted. © 1996 American Chemical Society

The adsorption of mixed nonionic/cationic surfactants at the air-water interface

Specular neutron reflection, in combination with hydrogen/deuterium isotopic substitution, has been used to investigate the adsorption of the mixed cationic-non-ionic surfactants hexadecyltrimethyl ammonium bromide, C16TAB,-hexaethylene glycol monododecyl ether, C12E6, at the air/water interface. Measurements were made at surfactant concentrations from below to above the critical micellar concentration, c.m.c., of the mixtures. For the C16TAB-C12E6 mixture, ideal mixing is observed at concentrations above the c.m.c. of the mixture whereas, below the c.m.c., departure from ideal mixing can be observed. The effects of temperature on the adsorption of the C16TAB-C12E6 mixture at the air/water interface are described. The total adsorption decreases with increasing temperature and the surface becomes richer in the non-ionic component of the mixture