Browsing by Author "Hoth, Markus"
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- ItemLight-Sheet Scattering Microscopy to Visualize Long-Term Interactions Between Cells and Extracellular Matrix(Lausanne : Frontiers Media, 2022) Zhou, Xiangda; Zhao, Renping; Yanamandra, Archana K.; Hoth, Markus; Qu, BinVisualizing 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.
- ItemOptoregulated force application to cellular receptors using molecular motors(London : Nature Publishing Group, 2021) Zheng, Yijun; Han, Mitchell K.L.; Zhao, Renping; Blass, Johanna; Zhang, Jingnan; Zhou, Dennis W.; Colard-Itté, Jean-Rémy; Dattler, Damien; Çolak, Arzu; Hoth, Markus; García, Andrés J.; Qu, Bin; Bennewitz, Roland; Giuseppone, Nicolas; del Campo, AránzazuProgress in our understanding of mechanotransduction events requires noninvasive methods for the manipulation of forces at molecular scale in physiological environments. Inspired by cellular mechanisms for force application (i.e. motor proteins pulling on cytoskeletal fibers), we present a unique molecular machine that can apply forces at cell-matrix and cell-cell junctions using light as an energy source. The key actuator is a light-driven rotatory molecular motor linked to polymer chains, which is intercalated between a membrane receptor and an engineered biointerface. The light-driven actuation of the molecular motor is converted in mechanical twisting of the entangled polymer chains, which will in turn effectively “pull” on engaged cell membrane receptors (e.g., integrins, T cell receptors) within the illuminated area. Applied forces have physiologically-relevant magnitude and occur at time scales within the relevant ranges for mechanotransduction at cell-friendly exposure conditions, as demonstrated in force-dependent focal adhesion maturation and T cell activation experiments. Our results reveal the potential of nanomotors for the manipulation of living cells at the molecular scale and demonstrate a functionality which at the moment cannot be achieved by other technologies for force application.
- ItemTargeting the Microtubule-Network Rescues CTL Killing Efficiency in Dense 3D Matrices(Lausanne : Frontiers Media, 2021) Zhao, Renping; Zhou, Xiangda; Khan, Essak S.; Alansary, Dalia; Friedmann, Kim S.; Yang, Wenjuan; Schwarz, Eva C.; Del Campo, Aránzazu; Hoth, Markus; Qu, BinEfficacy 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.
- ItemUnspecific CTL Killing Is Enhanced by High Glucose via TNF-Related Apoptosis-Inducing Ligand(Lausanne : Frontiers Media, 2022) Yang, Wenjuan; Denger, Andreas; Diener, Caroline; Küppers, Frederic; Soriano-Baguet, Leticia; Schäfer, Gertrud; Yanamandra, Archana K.; Zhao, Renping; Knörck, Arne; Schwarz, Eva C.; Hart, Martin; Lammert, Frank; Roma, Leticia Prates; Brenner, Dirk; Christidis, Grigorios; Helms, Volkhard; Meese, Eckart; Hoth, Markus; Qu, BinTNF-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.