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The erection of peripheral fences around protected areas has been shown to adversely affect the movement behavior, genetic connectivity and long-term viability of animal populations. However, fencing is also used extensively to reduce human encroachment, limit poaching activities, and mitigate human–wildlife conflicts. Limited information on the movement behavior of carnivores is available along the periphery of protected areas, particularly in semi-arid environments. To quantify the potential effects of the 824 km perimeter fence surrounding the Etosha National Park (Etosha, Namibia) on the movement and space use of carnivores, we collected GPS movement data from 36 lions (Panthera leo) and seven spotted hyenas (Crocuta crocuta) over a 14-year period. For these two apex carnivores, we measured the impact of the Etosha fence on their movements (step lengths and path straightness), calculated the likelihood of fence crossings, and evaluated the spatial–temporal characteristics of these crossings. Our results indicated that lions and hyenas moved faster (longer step lengths) when closer to the fence. Lions showed no variation in path straightness with respect to the fence but hyenas had straighter movement paths when closer to the fence. When moving within the vicinity of the fence, lions had a 9% likelihood of crossing the fence, while hyenas were much more likely (18%) to do so, which suggests that the Etosha fence was more permeable for hyenas than for lions. Fence crossings predominantly occurred at night for both species. Lions were more likely to cross the fence during the cold dry season (May–August) whereas hyenas crossed more often during the warm wet season (December–April). Female and adult lions had a higher probability of crossing the fence than males and subadults. These findings could be useful in the development of appropriate strategies to mitigate human–carnivore conflicts and thus promote the conservation of carnivores in multiple-use areas along the periphery of the park. Further research is needed to better elucidate the impact of perimeter fences on a broader suite of carnivores and other large mammals, including species-specific variation in the response to fences, as well as the economic benefits of fencing provided to local communities.

IntroductionMathematics classrooms are typically characterized by considerable heterogeneity with respect to students’ knowledge and skills. Mathematics teachers need to be highly attentive to students’ thinking, learning difficulties, and any misconceptions that they may develop. Identification of potential errors and appropriate ways to approach them is crucial for attaining positive learning outcomes. This paper explores which knowledge and affective-motivational skills teachers most require to effectively identify and approach students’ errors.MethodsTo address this research question within the German follow-up study of the Teacher Education and Development Study in Mathematics (TEDS-M), 131 primary school mathematics teachers’ ability to identify students’ errors was assessed based on (a) a digitalized speed test showing different students’ solutions in a written notation and (b) three video vignettes that showed different scenes from mathematics classes. These scenes dealt, among other things, with children who struggled with the lesson’s mathematical content. Teachers were asked to analyze students’ thinking and to determine how best to react. In addition, teachers’ mathematics pedagogical content knowledge, mathematical content knowledge, and beliefs were assessed in separate tests and served as predictors for teachers’ abilities to identify, analyze, and deal with students’ errors.ResultsThe results indicate that all components are interrelated. However, path analysis reveals that teachers’ ability to deal with students’ errors is mainly predicted by their constructivist beliefs while their ability to quickly identify typical students’ errors is largely dependent on their mathematics content knowledge.DiscussionThe results show the central filtering function of beliefs. Teachers who believe that students must shape and create their own learning processes are more successful in perceiving and analyzing student errors in classroom situations. They may understand errors as learning opportunities and - thus - pay specific attention to these occurrences.

Long-term human-environmental interactions in naturally fragile drylands are a focus of geomorphological and geoarchaeological research. Furthermore, many dryland societies were also affected by seismic activity. The semi-arid Shiraki Plain in the tectonically active southeastern Caucasus is currently covered by steppe and largely devoid of settlements. However, numerous Late Bronze to Early Iron Age city-type settlements suggest early state formation between ca. 3.2-2.5 ka that abruptly ended after that time. A paleolake was postulated for the lowest plain, and nearby pollen records suggest forest clearcutting of the upper altitudes under a more humid climate during the Late Bronze/Early Iron Ages. Furthermore, also an impact of earthquakes on regional Early Iron Age settlements was suggested. However, regional paleoenvironmental changes and paleoseismicity were not systematically studied so far. We combined geomorphological, sedimentological, chronological and paleoecological data with hydrological modelling to reconstruct regional Holocene paleoenvironmental changes, to identify natural and human causes and to study possible seismic events during the Late Bronze/Early Iron Ages. Our results show a balanced to negative Early to Mid-Holocene water balance probably caused by forested upper slopes. Hence, no lake but a pellic Vertisol developed in the lowest plain. Following, Late Bronze/Early Iron Age forest clear-cutting caused lake formation and the deposition of lacustrine sediments derived from soil erosion. Subsequently, regional aridification caused slow lake desiccation. Remains of freshwater fishes indicate that the lake potentially offered valuable ecosystem services for regional prehistoric societies even during the desiccation period. Finally, colluvial coverage of the lake sediments during the last centuries could have been linked with hydrological extremes during the Little Ice Age. Our study demonstrates that the Holocene hydrological balance of the Shiraki Plain was and is situated near a major hydrological threshold, making the landscape very sensitive to small-scale human or natural influences with severe consequences for local societies. Furthermore, seismites in the studied sediments do not indicate an influence of earthquakes on the main and late phases of Late Bronze/Early Iron Age settlement. Altogether, our study underlines the high value of multi-disciplinary approaches to investigate human-environmental interactions and paleoseismicity in drylands on millennial to centennial time scales.

Immunological memory is important to protect humans against recurring diseases. Memory CD8+ T cells are required for quick expansion into effector cells but also provide immediate cytotoxicity against their targets. Whereas many functions of the two main cytotoxic subtypes, effector memory CD8+ T cells (TEM) and central memory CD8+ T cells (TCM), are well defined, single TEM and TCM cell cytotoxicity has not been quantified. To quantify cytotoxic efficiency of TEM and TCM, we developed a FRET-based single cell fluorescent assay with NALM6 target cells which allows analysis of target cell apoptosis, secondary necrosis following apoptosis, and primary necrosis after TEM- or TCM-target cell contact. Both, single cell and population cytotoxicity assays reveal a higher cytotoxic efficiency of TEM compared to TCM, as quantified by target cell apoptosis and secondary necrosis. Perforin, granzyme B, FasL, but not TRAIL expression are higher in TEM compared to TCM. Higher perforin levels (likely in combination with higher granzyme levels) mediate higher cytotoxic efficiency of TEM compared to TCM. Both, TEM and TCM need the same time to find their targets, however contact time between CTL and target, time to induce apoptosis, and time to induce secondary necrosis are all shorter for TEM. In addition, immune synapse formation in TEM appears to be slightly more efficient than in TCM. Defining and quantifying single TEM and TCM cytotoxicity and the respective mechanisms is important to optimize future subset-based immune therapies.

TNF-related apoptosis inducing ligand (TRAIL) is expressed on cytotoxic T lymphocytes (CTLs) and TRAIL is linked to progression of diabetes. However, the impact of high glucose on TRAIL expression and its related killing function in CTLs still remains largely elusive. Here, we report that TRAIL is substantially up-regulated in CTLs in environments with high glucose (HG) both in vitro and in vivo. Non-mitochondrial reactive oxygen species, NFκB and PI3K/Akt are essential in HG-induced TRAIL upregulation in CTLs. TRAILhigh CTLs induce apoptosis of pancreatic beta cell line 1.4E7. Treatment with metformin and vitamin D reduces HG-enhanced expression of TRAIL in CTLs and coherently protects 1.4E7 cells from TRAIL-mediated apoptosis. Our work suggests that HG-induced TRAILhigh CTLs might contribute to the destruction of pancreatic beta cells in a hyperglycemia condition.

Visualizing interactions between cells and the extracellular matrix (ECM) mesh is important to understand cell behavior and regulatory mechanisms by the extracellular environment. However, long term visualization of three-dimensional (3D) matrix structures remains challenging mainly due to photobleaching or blind spots perpendicular to the imaging plane. Here, we combine label-free light-sheet scattering microcopy (LSSM) and fluorescence microscopy to solve these problems. We verified that LSSM can reliably visualize structures of collagen matrices from different origin including bovine, human and rat tail. The quality and intensity of collagen structure images acquired by LSSM did not decline with time. LSSM offers abundant wavelength choice to visualize matrix structures, maximizing combination possibilities with fluorescently-labelled cells, allowing visualizing of long-term ECM-cell interactions in 3D. Interestingly, we observed ultrathin thread-like structures between cells and matrix using LSSM, which were not observed by normal fluorescence microscopy. Transient local alignment of matrix by cell-applied forces can be observed. In summary, LSSM provides a powerful and robust approach to investigate the complex interplay between cells and ECM.

Efficacy of cytotoxic T lymphocyte (CTL)-based immunotherapy is still unsatisfactory against solid tumors, which are frequently characterized by condensed extracellular matrix. Here, using a unique 3D killing assay, we identify that the killing efficiency of primary human CTLs is substantially impaired in dense collagen matrices. Although the expression of cytotoxic proteins in CTLs remained intact in dense collagen, CTL motility was largely compromised. Using light-sheet microscopy, we found that persistence and velocity of CTL migration was influenced by the stiffness and porosity of the 3D matrix. Notably, 3D CTL velocity was strongly correlated with their nuclear deformability, which was enhanced by disruption of the microtubule network especially in dense matrices. Concomitantly, CTL migration, search efficiency, and killing efficiency in dense collagen were significantly increased in microtubule-perturbed CTLs. In addition, the chemotherapeutically used microtubule inhibitor vinblastine drastically enhanced CTL killing efficiency in dense collagen. Together, our findings suggest targeting the microtubule network as a promising strategy to enhance efficacy of CTL-based immunotherapy against solid tumors, especially stiff solid tumors.

The B-cell receptor (BCR) signaling pathway is a crucial pathway of B cells, both for their survival and for antigen-mediated activation, proliferation and differentiation. Its activation is also critical for the genesis of many lymphoma types. BCR-mediated lymphoma proliferation may be caused by activating BCR-pathway mutations and/or by active or tonic stimulation of the BCR. BCRs of lymphomas have frequently been described as polyreactive. In this review, the role of specific target antigens of the BCRs of lymphomas is highlighted. These antigens have been found to be restricted to specific lymphoma entities. The antigens can be of infectious origin, such as H. pylori in gastric MALT lymphoma or RpoC of M. catarrhalis in nodular lymphocyte predominant Hodgkin lymphoma, or they are autoantigens. Examples of such autoantigens are the BCR itself in chronic lymphocytic leukemia, LRPAP1 in mantle cell lymphoma, hyper-N-glycosylated SAMD14/neurabin-I in primary central nervous system lymphoma, hypo-phosphorylated ARS2 in diffuse large B-cell lymphoma, and hyper-phosphorylated SLP2, sumoylated HSP90 or saposin C in plasma cell dyscrasia. Notably, atypical posttranslational modifications are often responsible for the immunogenicity of many autoantigens. Possible therapeutic approaches evolving from these specific antigens are discussed.

Recently, neurabin-I and SAMD14 have been described as the autoantigenic target of approximately 66% of B-cell receptors (BCRs) of primary central nervous system lymphomas (PCNSL). Neurabin-I and SAMD14 share a highly homologous SAM domain that becomes immunogenic after atypical hyper-N-glycosylation (SAMD14 at ASN339 and neurabin-I at ASN1277). This post-translational modification of neurabin-I and SAMD14 seems to lead to a chronic immune reaction with B-cell receptor activation contributing to lymphoma genesis of PCNSLs. The selective tropism of PCNSL to the CNS corresponds well to the neurabin-I and SAMD14 protein expression pattern. When conjugated to Pseudomonas Exotoxin A (ETA´), the PCNSL reactive epitope exerts cytotoxic effects on lymphoma cells expressing a SAMD14/neurabin-I reactive BCR. Thus, the reactive epitopes of SAMD14/neurabin-I might be useful to establish additional therapeutic strategies against PCNSL. To test this possibility, we integrated the PCNSL-reactive epitope of SAMD14/neurabin-I into a heavy-chain-only Fab antibody format in substitution of the variable region. Specific binding of the prokaryotically produced SAMD14/neurabin-I Fab-antibody to lymphoma cells and their internalization were determined by flow cytometry. Since no established EBV-negative PCNSL cell line exists, we used the ABC-DLBCL cell lines OCI-Ly3 and U2932, which were transfected to express a SAMD14/neurabin-I reactive BCR. The SAMD14/neurabin-I Fab antibody bound specifically to DLBCL cells expressing a BCR with reactivity to SAMD14/neurabin-I and not to unmanipulated DLBCL cell lines. Eukaryotically produced full-length IgG antibodies are well established as immunotherapy format. Therefore, the PCNSL-reactive epitope of SAMD14/neurabin-I was cloned into a full-length IgG1 format replacing the variable domains of the light and heavy chains. The IgG1-format SAMD14/neurabin-I construct was found to specifically bind to target lymphoma cells expressing a SAMD14/neurabin-I reactive B cell receptor. In addition, it induced dose-dependent relative cytotoxicity against these lymphoma cells when incubated with PBMCs. Control DLBCL cells are not affected at any tested concentration. When integrated into the Fab-format and IgG1-format, the PCNSL-reactive epitope of SAMD14/neurabin-I functions as B-cell receptor Antigen for Reverse targeting (BAR). In particular, the IgG1-format BAR-body approach represents a very attractive therapeutic format for the treatment of PCNSLs, considering its specificity against SAMD14/neurabin-I reactive BCRs and the well-known pharmacodynamic properties of IgG antibodies.