Human germinal center B cells differ from naive and memory B cells by their aggregated MHC class II‐rich compartments lacking HLA‐DO

To generate memory B cells bearing high‐affinity antibodies, naive B cells first encounter antigen in the T cell‐rich areas of secondary lymphoid organs. There, they are activated by antigen‐specific T cells and become germinal center (GC) founder B cells. GC founders enter the GC to become centroblasts that proliferate and mutate their BCR. Centroblasts differentiate into centrocytes that undergo selection, which requires both the recognition/capture of antigen on follicular dendritic cells and the presentation of processed antigen to GC T cells. Because at each stage of differentiation B cells act as antigen‐presenting cells, we analyzed their content of HLA‐DR+‐rich compartments (MIIC), as well as their expression of HLA‐DM, which catalyzes peptide loading of class II molecules, and HLA‐DO, which interacts with HLA‐DM and focuses MHC class II peptide loading on antigens internalized by the BCR. Naive and memory B cells concentrate HLA‐DR, ‐DM and ‐DO into compartments dispersed under the cell surface, which are identified by their expression of lysosome‐associated membrane protein (Lamp)‐1 as late endosomes/lysosomes. GC founders and GC B cells express larger Lamp‐1+DR+ compartments that are concentrated in the juxta‐nuclear region. These compartments express lower levels of HLA‐DM and virtually no HLA‐DO. Upon induction of a GC founder phenotype through the prolonged (days) co‐ligation of BCR and CD40, the naive B cell's peripheral DR+DM+Lamp‐1+ compartments aggregate in a polar fashion close to the nucleus. Furthermore, HLA‐DO expression virtually disappears, whereas low levels of HLA‐DM remain co‐localized with HLA‐DR. Anti‐κ/λ antibodies, used as surrogate antigens, are promptly (minutes) endocytosed in naive, memory and GC B cells. Then, naive and memory B cells target the surrogate antigen to their peripheral HLA‐DO+ MIIC, while GC B cells target it to their HLA‐DO- MIIC aggregates. Taken together, our results show that human GC B cells differ from naive and memory B cells by their aggregated MIIC that lack HLA‐D

Innovating Pedagogy 2015: Open University Innovation Report 4

This series of reports explores new forms of teaching, learning and assessment for an interactive world, to guide teachers and policy makers in productive innovation. This fourth report proposes ten innovations that are already in currency but have not yet had a profound influence on education. To produce it, a group of academics at the Institute of Educational Technology in The Open University collaborated with researchers from the Center for Technology in Learning at SRI International. We proposed a long list of new educational terms, theories, and practices. We then pared these down to ten that have the potential to provoke major shifts in educational practice, particularly in post-school education. Lastly, we drew on published and unpublished writings to compile the ten sketches of new pedagogies that might transform education. These are summarised below in an approximate order of immediacy and timescale to widespread implementation

Interference of Distinct Invariant Chain Regions with Superantigen Contact Area and Antigenic Peptide Binding Groove of HLA-DR

In the endoplasmic reticulum, MHC class II ab dimers associate with the trimeric invariant chain (li), generating a nine-subunit(abli)3 complex. In the presence of li, the peptide binding groove is blocked, so that loading with self or antigenic peptides can only occur after proteolytic removal of li in specialized post-Golgi compartments. The class 11-associated invariant chain peptide region of li (about residues 81-1 04) is known to mediate binding to class II molecules and blockade of the groove, but this does not exclude additional contact sites for li. Using a set of overlapping li peptides and recombinant soluble li, we demonstrate here that a large segment of Ii encompassing approximately residues 71 to 128 interacts with HLA-DR molecules. The N- and C-terminal regions of this Ii segment appear to bind outside the peptide groove to the contact area for the staphylococcal superantigen Staphylococcus aureus enterotoxin B on the a1 domain. The core region of this segment (residues 95-108)prevents binding of antigenic peptides, probably by interaction with the peptide groove. Occupation of the groove with antigenic peptides abolishes binding not only of the core region, but also that of those Ii peptides that bind outside the groove. These findings suggest the existence of distinct conformational states of class II molecules, with Ii binding preferentially to one form

The Molecular Mechanism for Receptor-Stimulated Iron Release from the Plasma Iron Transport Protein Transferrin

Human transferrin receptor 1 (TfR) binds iron-loaded transferrin (Fe-Tf) and transports it to acidic endosomes where iron is released in a TfR-facilitated process. Consistent with our hypothesis that TfR binding stimulates iron release from Fe-Tf at acidic pH by stabilizing the apo-Tf conformation, a TfR mutant (W641A/F760A-TfR) that binds Fe-Tf, but not apo-Tf, cannot stimulate iron release from Fe-Tf, and less iron is released from Fe-Tf inside cells expressing W641A/F760A-TfR than cells expressing wild-type TfR (wtTfR). Electron paramagnetic resonance spectroscopy shows that binding at acidic pH to wtTfR, but not W641A/F760A-TfR, changes the Tf iron binding site ≥30 Å from the TfR W641/F760 patch. Mutation of Tf histidine residues predicted to interact with the W641/F760 patch eliminates TfR-dependent acceleration of iron release. Identification of TfR and Tf residues critical for TfR-facilitated iron release, yet distant from a Tf iron binding site, demonstrates that TfR transmits long-range conformational changes and stabilizes the conformation of apo-Tf to accelerate iron release from Fe-Tf

Structure Sensitivity of CO2Conversion over Nickel Metal Nanoparticles Explained by Micro-Kinetics Simulations

Nickel metal nanoparticles are intensively researched for the catalytic conversion of carbon dioxide. They are commercially explored in the so-called power-to-methane application in which renewably resourced H2 reacts with CO2 to produce CH4, which is better known as the Sabatier reaction. Previous work has shown that this reaction is structure-sensitive. For instance, Ni/SiO2 catalysts reveal a maximum performance when nickel metal nanoparticles of ∼2-3 nm are used. Particularly important to a better understanding of the structure sensitivity of the Sabatier reaction over nickel-based catalysts is to understand all relevant elementary reaction steps over various nickel metal facets because this will tell as to which type of nickel facets and which elementary reaction steps are crucial for designing an efficient nickel-based methanation catalyst. In this work, we have determined by density functional theory (DFT) calculations and micro-kinetics modeling (MKM) simulations that the two terrace facets Ni(111) and Ni(100) and the stepped facet Ni(211) barely show any activity in CO2 methanation. The stepped facet Ni(110) turned out to be the most effective in CO2 methanation. Herein, it was found that the dominant kinetic route corresponds to a combination of the carbide and formate reaction pathways. It was found that the dissociation of H2CO∗ toward CH2∗ and O∗ is the most critical elementary reaction step on this Ni(110) facet. The calculated activity of a range of Wulff-constructed nickel metal nanoparticles, accounting for varying ratios of the different facets and undercoordinated atoms exposed, reveals the same trend of activity-versus-nanoparticle size, as was observed in previous experimental work from our research group, thereby providing an explanation for the structure-sensitive nature of the Sabatier reaction

Structural and functional protein network analyses predict novel signaling functions for rhodopsin

Proteomic analyses, literature mining, and structural data were combined to generate an extensive signaling network linked to the visual G protein-coupled receptor rhodopsin. Network analysis suggests novel signaling routes to cytoskeleton dynamics and vesicular trafficking

Distinct immune stimulatory effects of anti-human VISTA antibodies are determined by Fc-receptor interaction

VISTA (PD-1H) is an immune regulatory molecule considered part of the next wave of immuno-oncology targets. VISTA is an immunoglobulin (Ig) superfamily cell surface molecule mainly expressed on myeloid cells, and to some extent on NK cells and T cells. In previous preclinical studies, some VISTA-targeting antibodies provided immune inhibitory signals, while other antibodies triggered immune stimulatory signals. Importantly, for therapeutic antibodies, the isotype backbone can have a strong impact on antibody function. To elucidate the mode of action of immune stimulatory anti-VISTA antibodies, we studied three different anti-human VISTA antibody clones, each on three different IgG isotypes currently used for therapeutic antibodies: unaltered IgG1 (IgG1-WT), IgG1-KO (IgG1-LL234,235AA-variant with reduced Fc-effector function), and IgG4-Pro (IgG4- S228P-variant with stabilized hinge region). Antibody functionality was analysed in mixed leukocyte reaction (MLR) of human peripheral blood mononuclear cells (PBMCs), as a model system for ongoing immune reactions, on unstimulated human PBMCs, as a model system for a resting immune system, and also on acute myeloid leukemia (AML) patient samples to evaluate anti-VISTA antibody effects on primary tumor material. The functions of three anti-human VISTA antibodies were determined by their IgG isotype backbones. An MLR of healthy donor PBMCs was effectively augmented by anti-VISTA-IgG4-Pro and anti-VISTA-IgG1-WT antibodies, as indicated by increased levels of cytokines, T cell activation markers and T cell proliferation. However, in a culture of unstimulated PBMCs of single healthy donors, only anti-VISTA-IgG1-WT antibodies increased the activation marker HLA-DR on resting myeloid cells, and chemokine levels. Interestingly, interactions with different Fc-receptors were required for these effects, namely CD64 for augmentation of MLR, and CD16 for activation of resting myeloid cells. Furthermore, anti-VISTA-IgG1-KO antibodies had nearly no impact in any model system. Similarly, in AML patient samples, anti-VISTA-antibody on IgG4-Pro backbone, but not on IgG1-KO backbone, increased interactions, as a novel readout of activity, between immune cells and CD34+ AML cancer cells. In conclusion, the immune stimulatory effects of antagonistic anti-VISTA antibodies are defined by the antibody isotype and interaction with different Fc-gamma-receptors, highlighting the importance of understanding these interactions when designing immune stimulatory antibody therapeutics for immuno-oncology applications

Whole grain-rich diet reduces body weight and systemic low-grade inflammation without inducing major changes of the gut microbiome: a randomised cross-over trial

Objective To investigate whether a whole grain diet alters the gut microbiome and insulin sensitivity, as well as biomarkers of metabolic health and gut functionality. Design 60 Danish adults at risk of developing metabolic syndrome were included in a randomised cross-over trial with two 8-week dietary intervention periods comprising whole grain diet and refined grain diet, separated by a washout period of ≥6 weeks. The response to the interventions on the gut microbiome composition and insulin sensitivity as well on measures of glucose and lipid metabolism, gut functionality, inflammatory markers, anthropometry and urine metabolomics were assessed. Results 50 participants completed both periods with a whole grain intake of 179±50 g/day and 13±10 g/day in the whole grain and refined grain period, respectively. Compliance was confirmed by a difference in plasma alkylresorcinols (p<0.0001). Compared with refined grain, whole grain did not significantly alter glucose homeostasis and did not induce major changes in the faecal microbiome. Also, breath hydrogen levels, plasma short-chain fatty acids, intestinal integrity and intestinal transit time were not affected. The whole grain diet did, however, compared with the refined grain diet, decrease body weight (p<0.0001), serum inflammatory markers, interleukin (IL)-6 (p=0.009) and C-reactive protein (p=0.003). The reduction in body weight was consistent with a reduction in energy intake, and IL-6 reduction was associated with the amount of whole grain consumed, in particular with intake of rye. Conclusion Compared with refined grain diet, whole grain diet did not alter insulin sensitivity and gut microbiome but reduced body weight and systemic low-grade inflammation