Browsing by Author "Del Campo, Aránzazu"
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- ItemInhibition of Collagenase Q1 of Bacillus cereus as a Novel Antivirulence Strategy for the Treatment of Skin-Wound Infections(Weinheim : Wiley-VCH Verlag, 2022) Alhayek, Alaa; Khan, Essak S.; Schönauer, Esther; Däinghaus, Tobias; Shafiei, Roya; Voos, Katrin; Han, Mitchell K. L.; Ducho, Christian; Posselt, Gernot; Wessler, Silja; Brandstetter, Hans; Haupenthal, Jörg; Del Campo, Aránzazu; Hirsch, Anna K. H.Despite the progress in surgical techniques and antibiotic prophylaxis, opportunistic wound infections with Bacillus cereus remain a public health problem. Secreted toxins are one of the main factors contributing to B. cereus pathogenicity. A promising strategy to treat such infections is to target these toxins and not the bacteria. Although the exoenzymes produced by B. cereus are thoroughly investigated, little is known about the role of B. cereus collagenases in wound infections. In this report, the collagenolytic activity of secreted collagenases (Col) is characterized in the B. cereus culture supernatant (csn) and its isolated recombinantly produced ColQ1 is characterized. The data reveals that ColQ1 causes damage on dermal collagen (COL). This results in gaps in the tissue, which might facilitate the spread of bacteria. The importance of B. cereus collagenases is also demonstrated in disease promotion using two inhibitors. Compound 2 shows high efficacy in peptidolytic, gelatinolytic, and COL degradation assays. It also preserves the fibrillar COLs in skin tissue challenged with ColQ1, as well as the viability of skin cells treated with B. cereus csn. A Galleria mellonella model highlights the significance of collagenase inhibition in vivo.
- ItemPhotoactivatable Hsp47: A tool to control and regulate collagen secretion & assembly(Hoboken, NJ : Wiley, 2018) Khan, Essak; Sankaran, Shrikrishnan; Paez, Julieta; Muth, Christina; Han, Mitchell; Del Campo, AránzazuCollagen is the most abundant structural protein in mammals and is crucial for the mechanical integrity of tissues. Hsp47, an endoplasmic reticulum resident collagen-specific chaperone, is involved in collagen biosynthesis and plays a fundamental role in the folding, stability, and intracellular transport of procollagen triple helices. This work reports on a photoactivatable derivative of Hsp47 that allows regulation of collagen biosynthesis within mammalian cells using light. Photoactivatable Hsp47 contains a non-natural light-responsive tyrosine (o-nitro benzyl tyrosine (ONBY)) at Tyr383 position of the protein sequence. This mutation renders Hsp47 inactive toward collagen binding. The inactive, photoactivatable protein is easily uptaken by cells within a few minutes of incubation, and accumulated at the endoplasmic reticulum via retrograde KDEL receptor-mediated uptake. Upon light exposure, the photoactivatable Hsp47 turns into functional Hsp47 in situ. The increased intracellular concentration of Hsp47 results in stimulated secretion of collagen. The ability to promote collagen synthesis on demand, with spatiotemporal resolution, and in diseased state cells is demonstrated in vitro. It is envisioned that photoactivatable Hsp47 allows unprecedented fundamental studies of collagen biosynthesis, matrix biology, and inspires new therapeutic concepts in biomedicine and tissue regeneration.
- 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.
- ItemToward light‐regulated living biomaterials(Hoboken, NJ : Wiley, 2018) Sankaran, Shrikrishnan; Zhao, Shifang; Muth, Christina; Paez, Julieta; Del Campo, AránzazuLiving materials are an emergent material class, infused with the productive,adaptive, and regenerative properties of living organisms. Property regulation in living materials requires encoding responsive units in the living components to allow external manipulation of their function. Here, an optoregulated Escherichia coli (E. coli)-based living biomaterial that can be externally addressed using light to interact with mammalian cells is demonstrated. This is achieved by using a photoactivatable inducer of gene expression and bacterial surface display technology to present an integrin-specific miniprotein on the outer membrane of an endotoxin-free E. coli strain. Hydrogel surfaces functionalized with the bacteria can expose cell adhesive molecules upon in situ light-activation, and trigger cell adhesion. Surface immobilized bacteria are able to deliver a fluorescent protein to the mammalian cells with which they are interacting, indicating the potential of such a bacterial material to deliver molecules to cells in a targeted manner.