Chicken γδ T cells proliferate upon IL-2 and IL-12 treatment and show a restricted receptor repertoire in cell culture

In chickens, γδ T cells represent a large fraction of peripheral T cells; however, their function remains largely unknown. Here, we describe the selective in vitro expansion of γδ T cells from total splenocytes by stimulation with the cytokines IL-2 and IL-12. Under these conditions, γδ T cells proliferated preferentially and reached frequencies of >95% within three weeks. Although IL-2 alone also triggered proliferation, an increased proliferation rate was observed in combination with IL-12. Most of the expanded cells were γδ TCR and CD8 double-positive. Splenocytes sorted into TCR1+CD8+, TCR1highCD8−, and TCR1lowCD8− subsets proliferated well upon dual stimulation with IL-2/IL-12, indicating that none of the three γδ T cell subsets require bystander activation for proliferation. TCR1+CD8+ cells maintained CD8 surface expression during stimulation, whereas CD8− subpopulations showed varied levels of CD8 upregulation, with the highest upregulation observed in the TCR1high subset. Changes in the γδ T-cell receptor repertoire during cell culture from day 0 to day 21 were analyzed by next-generation sequencing of the γδ variable regions. Overall, long-term culture led to a restricted γ and δ chain repertoire, characterized by a reduced number of unique variable region clonotypes, and specific V genes were enriched at day 21. On day 0, the δ chain repertoire was highly diverse, and the predominant clonotypes differed between animals, while the most frequent γ-chain clonotypes were shared between animals. However, on day 21, the most frequent clonotypes in both the γ and δ chain repertoires were different between animals, indicating that selective expansion of dominant clonotypes during stimulation seems to be an individual outcome. In conclusion, IL-2 and IL-12 were sufficient to stimulate the in vitro outgrowth of γδ T cells. Analyses of the TCR repertoire indicate that the culture leads to an expansion of individual T cell clones, which may reflect previous in vivo activation. This system will be instrumental in studying γδ T cell function

Insights into intermetallic phases on pulse welded dissimilar metal joints

The Magnetic Pulse Welding (MPW) process has been developed to an industrially used joining method which is considered to be a fast, noncontact, clean and "cold" solid state welding process. Unlike fusion welding, the absence of direct heat during the welding cycle makes it possible to join dissimilar metals, for instance aluminium to copper or copper to steel, without noticeable detrimental metallurgical defects. This is very desirable, as today s industry lacks technologies to join often not fusion-weldable dissimilar materials effectively. However, current metallographic studies show that for many material combinations the formation of intermetallic seams in the joint region of magnetic pulse welds can not be completely avoided. Modern technical equipment for MPW is used to join aluminium with copper in order to study the microstructure and the intermetallic phases formed in the weld region in dependence of the processing parameters. The welds are analysed by means of metallographic and electron microscopic (SEM) methods. Relations between the parameters and the microstructures formed within the weld joints are shown. Based on the obtained results conclusions will be drawn with respect to the intermetallic phase formation process and the optimization of the weld microstructure and properties

A Latent Class Analysis of University Lecturers’ Switch to Online Teaching during the First COVID-19 Lockdown: The Role of Educational Technology, Self-Efficacy, and Institutional Support

The switch to emergency remote teaching (ERT) due to the first COVID-19 lockdown demanded a lot from university lecturers yet did not pose the same challenge to all of them. This study sought to explain differences among lecturers (n = 796) from universities in France, Germany, Switzerland, and the UK in their use of educational technology for teaching, institutional support, and personal factors. Guided by the Social Cognitive Theory (SCT), lecturers’ behavior (educational technology use), environment (institutional support), and personal factors (ERT self-efficacy, continuance intentions, and demographics) were examined. Latent class analysis was employed to identify different types of lecturers in view of educational technology use, while multinomial regression and Wald chi-square test were used to distinguish classes. The largest latent class were Presenters (45.6%), who focused on content delivery, followed by Strivers (22.1%), who strived for social interaction, Routineers (19.6%), who were ready for online teaching, and Evaders (12.7%), who evaded using technology for educational purposes. Both personal factors and perceived institutional support explained class membership significantly. Accordingly, Evaders were older, less experienced, and rarely perceived institutional support as useful. Routineers, the Evaders’ counterparts, felt most self-efficient in ERT and held the highest continuance intentions for educational technology use. This research suggests that universities engage lecturers in evidence-based professional development that seeks shared visions of digital transformation, networks and communities, and design-based researc

Characterization of a novel chicken γδ TCR-specific marker

Chickens are a species with a high number of γδ T cells in various tissues. Despite their abundance, γδ T cells are poorly characterized in chickens, partially due to a lack of specific reagents to characterize these cells. Up until now, the TCR1 clone has been the only γδ T cell-specific monoclonal antibody (mAb) in chickens and additional reagents for γδ T cell subsets are needed. In order to address this issue, new mAb were generated in our laboratory by immunizing mice with in vitro cultured γδ T cells. In an initial flow cytometric screen a new mAb, clone “8D2”, displayed an interesting staining pattern that mirrored γδ TCR up- and downregulation in the γδ T cell line D4 over time, prompting us to characterize this antibody further. We compared the expression of the unknown 8D2 epitope in combination with TCR1 staining across various primary cells. In splenocytes, peripheral blood lymphocytes and intestinal epithelial cells, 8D2 consistently labeled a subset of TCR1+ cells. To determine, whether specific γδ T cell receptors were recognized by 8D2, we sorted γδ T cells according to their 8D2 and TCR1 expression and analyzed their TCR V(D)J gene usage by TCR profiling. Strikingly, sorted 8D2+ cells preferentially expressed Vγ3 genes, whereas the TCR Vγ genes used by TCR1+ 8D2- cells were more variable. γδ TCR in 8D2+ cells were most frequently comprised of gamma chain VJ genes TRGV3-8 and TRGJ3, and delta chain VDJ genes TRDV1-2, TRDD2, TRDJ1. To confirm binding of 8D2 to specific γδ TCR, the preferentially utilized combination of TRG and TRD was expressed in HEK293 cells in combination with CD3, demonstrating surface binding of the 8D2 mAb to this Vγ3 γδ TCR-expressing cell line. Conversely, HEK293 cells expressing either Vγ1 or Vγ2 TCR did not react with 8D2. In conclusion, 8D2 is a novel tool for identifying specific Vγ3 bearing γδ T cells

Influence of Axial Workpiece Positioning during Magnetic Pulse Welding of Aluminum-Steel Joints

Magnetic Pulse Welding (MPW) offers a method to economically join similar and dissimilar metals without the need for external physical or chemical binders, while avoiding the adverse heating effects seen in many welding techniques. MPW allows for the fabrication of joints via the harnessing of Lorentz forces, which result from discharging a current pulse through a coil. In the process an outer piece (flyer) is accelerated onto an inner piece (parent), and welding is achieved using propagating impact fronts. There are several geometrical factors to be considered including the flyer-coil distance, the parentflyer distance, as well as the axial relationship between flyer and coil (working length). Various shapes of the front are possible and each configuration has its own advantages and drawbacks. The goal of this work is to show not only how the aforementioned parameters are related, but also ways to optimize front propagations, which are vital to the welding result. This is done primarily by determining the influence of the working length of tubular MPW specimens. It is shown that for steel-aluminum joints in the given arrangements, three different front regimes exist, which are related to geometrical factors. These results are especially useful to avoid seemingly favorable but nevertheless suboptimal conditions for flyer movement that would reduce weld quality and energy efficiency of the process

Interleukin-18 Mediates Immune Responses to Campylobacter jejuni Infection in Gnotobiotic Mice

Background Human Campylobacter jejuni infections are progressively rising worldwide. Information about the molecular mechanisms underlying campylobacteriosis, however, are limited. In the present study we investigated whether cytokines such as IL-23, IL-22 and IL-18, which share pivotal functions in host immunity, were involved in mediating intestinal and systemic immunopathological responses upon C. jejuni infection. Methodology/Principal Findings To assure stable infection, gnotobiotic (i.e. secondary abiotic) IL- 23p19-/-, IL-22-/- and IL-18-/- mice were generated by broad-spectrum antibiotic treatment. Following peroral C. jejuni strain 81–176 infection, mice of all genotypes harbored comparably high pathogenic loads in their intestines. As compared to wildtype controls, however, IL-18-/- mice displayed less distinct C. jejuni induced sequelae as indicated by less pronounced large intestinal shrinkage and lower numbers of apoptotic cells in the colonic epithelial layer at day 8 postinfection (p.i.). Furthermore, lower colonic numbers of adaptive immune cells including regulatory T cells and B lymphocytes were accompanied by less distinct secretion of pro-inflammatory cytokines such as TNF and IFN-γ and lower IL-17A mRNA expression levels in colonic ex vivo biopsies of infected IL-18-/- as compared to wildtype mice. Upon C. jejuni infection, colonic IL-23p19 expression was up-regulated in IL-18-/- mice only, whereas IL-22 mRNA levels were lower in uninfected and infected IL-23p19-/- as well as infected IL-18-/- as compared to respective wildtype control mice. Remarkably, not only intestinal, but also systemic infection-induced immune responses were less pronounced in IL-18-/- mice as indicated by lower TNF, IFN-γ and IL-6 serum levels as compared to wildtype mice. Conclusion/Significance We here show for the first time that IL-18 is essentially involved in mediating C. jejuni infection in the gnotobiotic mouse model. Future studies need to further unravel the underlying regulatory mechanisms orchestrating pathogen-host interaction