Affinity-restricted memory B cells dominate recall responses to heterologous flaviviruses

Memory B cells (MBCs) can respond to heterologous antigens either by molding new specificities through secondary germinal centers (GCs) or selecting pre-existing clones without further affinity maturation. To distinguish these mechanisms in flavivirus infections and immunizations, we studied recall responses to envelope protein domain III (DIII). Conditional deletion of activation induced cytidine deaminase (AID) between heterologous challenges of West Nile, Japanese encephalitis, Zika, and Dengue viruses did not affect recall responses. DIII-specific MBCs were contained mostly within the plasma cell-biased CD80(+) subset and few GCs arose following heterologous boosters, demonstrating that recall responses are confined by pre-existing clonal diversity. Measurement of monoclonal antibody binding affinity to DIII proteins, timed AID deletion, single cell RNA-sequencing, and lineage tracing experiments point to selection of relatively low affinity MBCs as a mechanism to promote diversity. Engineering immunogens to avoid this MBC diversity may facilitate flavivirus type-specific vaccines with minimized potential for infection enhancement

pO polarography, contrast enhanced color duplex sonography (CDS), [18F] fluoromisonidazole and [18F] fluorodeoxyglucose positron emission tomography: validated methods for the evaluation of therapy-relevant tumor oxygenation or only bricks in the puzzle of tumor hypoxia?

<p>Abstract</p> <p>Background</p> <p>The present study was conducted to analyze the value of ([¹⁸F] fluoromisonidazole (FMISO) and [¹⁸F]-2-fluoro-2'-deoxyglucose (FDG) PET as well as color pixel density (CPD) and tumor perfusion (TP) assessed by color duplex sonography (CDS) for determination of therapeutic relevant hypoxia. As a standard for measuring tissue oxygenation in human tumors, the invasive, computerized polarographic needle electrode system (pO₂histography) was used for comparing the different non invasive measurements.</p> <p>Methods</p> <p>Until now a total of 38 Patients with malignancies of the head and neck were examined. Tumor tissue pO₂was measured using a pO₂-histograph. The needle electrode was placed CT-controlled in the tumor without general or local anesthesia. To assess the biological and clinical relevance of oxygenation measurement, the relative frequency of pO₂readings, with values ≤ 2.5, ≤ 5.0 and ≤ 10.0 mmHg, as well as mean and median pO₂were stated. FMISO PET consisted of one static scan of the relevant region, performed 120 min after intravenous administration. FMISO tumor to muscle ratios (FMISO_T/M) and tumor to blood ratios (FMISO_T/B) were calculated. FDG PET of the lymph node metastases was performed 71 ± 17 min after intravenous administration. To visualize as many vessels as possible by CDS, a contrast enhancer (Levovist^®, Schering Corp., Germany) was administered. Color pixel density (CPD) was defined as the ratio of colored to grey pixels in a region of interest. From CDS signals two parameters were extracted: color hue – defining velocity (v) and color area – defining perfused area (A). Signal intensity as a measure of tissue perfusion (TP) was quantified as follows: TP = v_mean× A_mean.</p> <p>Results</p> <p>In order to investigate the degree of linear association, we calculated the Pearson correlation coefficient. Slight (|r| > 0.4) to moderate (|r| > 0.6) correlation was found between the parameters of pO₂polarography (pO₂readings with values ≤ 2.5, ≤ 5.0 and ≤ 10.0 mmHg, as well as median pO₂), CPD and FMISO_T/M. Only a slight correlation between TP and the fraction of pO₂values ≤ 10.0 mmHg, median and mean pO₂could be detected. After exclusion of four outliers the absolute values of the Pearson correlation coefficients increased clearly. There was no relevant association between mean or maximum FDG uptake and the different polarographic- as well as the CDS parameters.</p> <p>Conclusion</p> <p>CDS and FMISO PET represent different approaches for estimation of therapy relevant tumor hypoxia. Each of these approaches is methodologically limited, making evaluation of clinical potential in prospective studies necessary.</p

Toward a theory of repeat purchase drivers for consumer services

The marketing discipline’s knowledge about the drivers of service customers’ repeat purchase behavior is highly fragmented. This research attempts to overcome that fragmented state of knowledge by making major advances toward a theory of repeat purchase drivers for consumer services. Drawing on means–end theory, the authors develop a hierarchical classification scheme that organizes repeat purchase drivers into an integrative and comprehensive framework. They then identify drivers on the basis of 188 face-to-face laddering interviews in two countries (USA and Germany) and assess the drivers’ importance and interrelations through a national probability sample survey of 618 service customers. In addition to presenting an exhaustive and coherent set of hierarchical repeat-purchase drivers, the authors provide theoretical explanations for how and why drivers relate to one another and to repeat purchase behavior. This research also tests the boundary conditions of the proposed framework by accounting for different service types. In addition to its theoretical contribution, the framework provides companies with specific information about how to manage long-term customer relationships successfully

Customer Interaction and Innovation in Hybrid Offerings:Investigating Moderation and Mediation Effects for Goods and Services Innovation

Hybrid offerings are bundles of goods and services offerings provided by the same firm. Bundling value offerings affects how firms innovate, interact with customers, and customize their goods and services. However, it remains unclear how customer interaction might drive the innovation performance of various bundled components. Therefore, this study investigates the effects of customer interactions and service customization on both goods and services innovations in a hybrid offering context, using a unique data set of 146 information technology and manufacturing firms. Customer interaction appears beneficial to both goods and services innovation in a hybrid offerings context, but service customization has different direct effects on goods versus services innovation. As a potential mediator, customer knowledge mobilization resources exert different effects on the goods and services elements of hybrid offerings. Furthermore, for high-interaction customers, medium levels of technical modularity lead to most favorable innovation outcomes for services innovation. The results thus suggest that providers of hybrid offerings should foster customer interactions, to drive the innovation performance of the good and service components, while still making sure to implement service customization strategies. These findings have notable implications for service innovation research

Structure-bias relationships for fenoterol stereoisomers in six molecular and cellular assays at the β2-adrenoceptor.

Functional selectivity is well established as an underlying concept of ligand-specific signaling via G protein-coupled receptors (GPCRs). Functionally, selective drugs could show greater therapeutic efficacy and fewer adverse effects. Dual coupling of the β2-adrenoceptor (β2AR) triggers a signal transduction via Gsα and Giα proteins. Here, we examined 12 fenoterol stereoisomers in six molecular and cellular assays. Using β2AR-Gsα and β2AR-Giα fusion proteins, (R,S')- and (S,S')-isomers of 4'-methoxy-1-naphthyl-fenoterol were identified as biased ligands with preference for Gs. G protein-independent signaling via β-arrestin-2 was disfavored by these ligands. Isolated human neutrophils constituted an ex vivo model of β2AR signaling and demonstrated functional selectivity through the dissociation of cAMP accumulation and the inhibition of formyl peptide-stimulated production of reactive oxygen species. Ligand bias was calculated using an operational model of agonism and revealed that the fenoterol scaffold constitutes a promising lead structure for the development of Gs-biased β2AR agonists

Dissociations in the effects of beta2-adrenergic receptor agonists on cAMP formation and superoxide production in human neutrophils: Support for the concept of functional selectivity

In neutrophils, activation of the beta2-adrenergic receptor (beta2AR), a Gs-coupled receptor, inhibits inflammatory responses, which could be therapeutically exploited. The aim of this study was to evaluate the effects of various beta2AR ligands on adenosine-3',5'-cyclic monophosphate (cAMP) accumulation and N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP)-induced superoxide anion (O2*-) production in human neutrophils and to probe the concept of ligand-specific receptor conformations (also referred to as functional selectivity or biased signaling) in a native cell system. cAMP concentration was determined by HPLC/tandem mass spectrometry, and O2*- formation was assessed by superoxide dismutase-inhibitable reduction of ferricytochrome c. beta2AR agonists were generally more potent in inhibiting fMLP-induced O2*- production than in stimulating cAMP accumulation. (-)-Ephedrine and dichloroisoproterenol were devoid of any agonistic activity in the cAMP assay, but partially inhibited fMLP-induced O2*- production. Moreover, (-)-adrenaline was equiefficacious in both assays whereas the efficacy of salbutamol was more than two-fold higher in the O2*- assay. In contrast to the agonists, the effects of beta2AR antagonists were comparable between the two parameters on neutrophils. Differences between the data from neutrophils and recombinant test systems were observed for the beta2AR agonists as well as for the beta2AR antagonists. Lastly, we obtained no evidence for an involvement of protein kinase A in the inhibition of fMLP-induced O2*- production after beta2AR-stimulation, although, in principle, cAMP-increasing substances can inhibit O2*- production. Taken together, our data corroborate the concept of ligand-specific receptor conformations with unique signaling capabilities and suggest that the beta2AR inhibits O2*- production in a cAMP-independent manner