@article{KarlWussmannKressetal.2019, author = {Karl, Franziska and Wußmann, Maximiliane and Kreß, Luisa and Malzacher, Tobias and Fey, Phillip and Groeber-Becker, Florian and {\"U}{\c{c}}eyler, Nurcan}, title = {Patient-derived in vitro skin models for investigation of small fiber pathology}, series = {Annals of Clinical and Translational Neurology}, volume = {6}, journal = {Annals of Clinical and Translational Neurology}, number = {9}, doi = {10.1002/acn3.50871}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-201649}, pages = {1797-1806}, year = {2019}, abstract = {Objective To establish individually expandable primary fibroblast and keratinocyte cultures from 3-mm skin punch biopsies for patient-derived in vitro skin models to investigate of small fiber pathology. Methods We obtained 6-mm skin punch biopsies from the calf of two patients with small fiber neuropathy (SFN) and two healthy controls. One half (3 mm) was used for diagnostic intraepidermal nerve fiber density (IENFD). From the second half, we isolated and cultured fibroblasts and keratinocytes. Cells were used to generate patient-derived full-thickness three-dimensional (3D) skin models containing a dermal and epidermal component. Cells and skin models were characterized morphologically, immunocyto- and -histochemically (vimentin, cytokeratin (CK)-10, CK 14, ki67, collagen1, and procollagen), and by electrical impedance. Results Distal IENFD was reduced in the SFN patients (2 fibers/mm each), while IENFD was normal in the controls (8 fibers/mm, 7 fibers/mm). Two-dimensional (2D) cultured skin cells showed normal morphology, adequate viability, and proliferation, and expressed cell-specific markers without relevant difference between SFN patient and healthy control. Using 2D cultured fibroblasts and keratinocytes, we obtained subject-derived 3D skin models. Morphology of the 3D model was analogous to the respective skin biopsy specimens. Both, the dermal and the epidermal layer carried cell-specific markers and showed a homogenous expression of extracellular matrix proteins. Interpretation Our protocol allows the generation of disease-specific 2D and 3D skin models, which can be used to investigate the cross-talk between skin cells and sensory neurons in small fiber pathology.}, language = {en} } @article{WallstabeBussemerGroeberBeckeretal.2020, author = {Wallstabe, Julia and Bussemer, Lydia and Groeber-Becker, Florian and Freund, Lukas and Alb, Mirian and Dragan, Mariola and Waaga-Gasser, Ana Maria and Jakubietz, Rafael and Kneitz, Hermann and Rosenwald, Andreas and Rebhan, Silke and Walles, Heike and Mielke, Stephan}, title = {Inflammation-Induced Tissue Damage Mimicking GvHD in Human Skin Models as Test Platform for Immunotherapeutics}, series = {ALTEX}, volume = {37}, journal = {ALTEX}, number = {3}, doi = {10.14573/altex.1907181}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-229974}, pages = {429-440}, year = {2020}, abstract = {Due to the rapidly increasing development and use of cellular products, there is a rising demand for non-animal-based test platforms to predict, study and treat undesired immunity. Here, we generated human organotypic skin models from human biopsies by isolating and expanding keratinocytes, fibroblasts and microvascular endothelial cells and seeding these components on a collagen matrix or a biological vascularized scaffold matrix in a bioreactor. We then were able to induce inflammation-mediated tissue damage by adding pre-stimulated, mismatched allogeneic lymphocytes and/or inflammatory cytokine-containing supernatants histomorphologically mimicking severe graft versus host disease (GvHD) of the skin. This could be prevented by the addition of immunosuppressants to the models. Consequently, these models harbor a promising potential to serve as a test platform for the prediction, prevention and treatment of GvHD. They also allow functional studies of immune effectors and suppressors including but not limited to allodepleted lymphocytes, gamma-delta T cells, regulatory T cells and mesenchymal stromal cells, which would otherwise be limited to animal models. Thus, the current test platform, developed with the limitation that no professional antigen presenting cells are in place, could greatly reduce animal testing for investigation of novel immune therapies.}, language = {en} } @article{GenslerLeikeimMoellmannetal.2020, author = {Gensler, Marius and Leikeim, Anna and M{\"o}llmann, Marc and Komma, Miriam and Heid, Susanne and M{\"u}ller, Claudia and Boccaccini, Aldo R. and Salehi, Sahar and Groeber-Becker, Florian and Hansmann, Jan}, title = {3D printing of bioreactors in tissue engineering: A generalised approach}, series = {PLoS One}, volume = {15}, journal = {PLoS One}, number = {11}, doi = {10.1371/journal.pone.0242615}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-231368}, year = {2020}, abstract = {3D printing is a rapidly evolving field for biological (bioprinting) and non-biological applications. Due to a high degree of freedom for geometrical parameters in 3D printing, prototype printing of bioreactors is a promising approach in the field of Tissue Engineering. The variety of printers, materials, printing parameters and device settings is difficult to overview both for beginners as well as for most professionals. In order to address this problem, we designed a guidance including test bodies to elucidate the real printing performance for a given printer system. Therefore, performance parameters such as accuracy or mechanical stability of the test bodies are systematically analysed. Moreover, post processing steps such as sterilisation or cleaning are considered in the test procedure. The guidance presented here is also applicable to optimise the printer settings for a given printer device. As proof of concept, we compared fused filament fabrication, stereolithography and selective laser sintering as the three most used printing methods. We determined fused filament fabrication printing as the most economical solution, while stereolithography is most accurate and features the highest surface quality. Finally, we tested the applicability of our guidance by identifying a printer solution to manufacture a complex bioreactor for a perfused tissue construct. Due to its design, the manufacture via subtractive mechanical methods would be 21-fold more expensive than additive manufacturing and therefore, would result in three times the number of parts to be assembled subsequently. Using this bioreactor we showed a successful 14-day-culture of a biofabricated collagen-based tissue construct containing human dermal fibroblasts as the stromal part and a perfusable central channel with human microvascular endothelial cells. Our study indicates how the full potential of biofabrication can be exploited, as most printed tissues exhibit individual shapes and require storage under physiological conditions, after the bioprinting process.}, language = {en} } @article{SchneiderKruseBernardellideMattosetal.2021, author = {Schneider, Verena and Kruse, Daniel and Bernardelli de Mattos, Ives and Z{\"o}phel, Saskia and Tiltmann, Kendra-Kathrin and Reigl, Amelie and Khan, Sarah and Funk, Martin and Bodenschatz, Karl and Groeber-Becker, Florian}, title = {A 3D in vitro model for burn wounds: monitoring of regeneration on the epidermal level}, series = {Biomedicines}, volume = {9}, journal = {Biomedicines}, number = {9}, issn = {2227-9059}, doi = {10.3390/biomedicines9091153}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-246068}, year = {2021}, abstract = {Burns affect millions every year and a model to mimic the pathophysiology of such injuries in detail is required to better understand regeneration. The current gold standard for studying burn wounds are animal models, which are under criticism due to ethical considerations and a limited predictiveness. Here, we present a three-dimensional burn model, based on an open-source model, to monitor wound healing on the epidermal level. Skin equivalents were burned, using a preheated metal cylinder. The healing process was monitored regarding histomorphology, metabolic changes, inflammatory response and reepithelialization for 14 days. During this time, the wound size decreased from 25\% to 5\% of the model area and the inflammatory response (IL-1β, IL-6 and IL-8) showed a comparable course to wounding and healing in vivo. Additionally, the topical application of 5\% dexpanthenol enhanced tissue morphology and the number of proliferative keratinocytes in the newly formed epidermis, but did not influence the overall reepithelialization rate. In summary, the model showed a comparable healing process to in vivo, and thus, offers the opportunity to better understand the physiology of thermal burn wound healing on the keratinocyte level.}, language = {en} } @article{TucaBernardellideMattosFunketal.2022, author = {Tuca, Alexandru-Cristian and Bernardelli de Mattos, Ives and Funk, Martin and Winter, Raimund and Palackic, Alen and Groeber-Becker, Florian and Kruse, Daniel and Kukla, Fabian and Lemarchand, Thomas and Kamolz, Lars-Peter}, title = {Orchestrating the dermal/epidermal tissue ratio during wound healing by controlling the moisture content}, series = {Biomedicines}, volume = {10}, journal = {Biomedicines}, number = {6}, issn = {2227-9059}, doi = {10.3390/biomedicines10061286}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-275115}, year = {2022}, abstract = {A balanced and moist wound environment and surface increases the effect of various growth factors, cytokines, and chemokines, stimulating cell growth and wound healing. Considering this fact, we tested in vitro and in vivo water evaporation rates from the cellulose dressing epicite\(^{hydro}\) when combined with different secondary dressings as well as the resulting wound healing efficacy in a porcine donor site model. The aim of this study was to evaluate how the different rates of water evaporation affected wound healing efficacy. To this end, epicite\(^{hydro}\) primary dressing, in combination with different secondary dressing materials (cotton gauze, JELONET\(^◊\), AQUACEL\(^®\) Extra\(^™\), and OPSITE\(^◊\) Flexifix), was placed on 3 × 3 cm-sized dermatome wounds with a depth of 1.2 mm on the flanks of domestic pigs. The healing process was analyzed histologically and quantified by morphometry. High water evaporation rates by using the correct secondary dressing, such as cotton gauze, favored a better re-epithelialization in comparison with the low water evaporation resulting from an occlusive secondary dressing, which favored the formation of a new and intact dermal tissue that nearly fully replaced all the dermis that was removed during wounding. This newly available evidence may be of great benefit to clinical wound management.}, language = {en} } @article{WeigelMalkmusWeigeletal.2022, author = {Weigel, Tobias and Malkmus, Christoph and Weigel, Verena and Wußmann, Maximiliane and Berger, Constantin and Brennecke, Julian and Groeber-Becker, Florian and Hansmann, Jan}, title = {Fully Synthetic 3D Fibrous Scaffolds for Stromal Tissues—Replacement of Animal-Derived Scaffold Materials Demonstrated by Multilayered Skin}, series = {Advanced Materials}, volume = {34}, journal = {Advanced Materials}, number = {10}, doi = {10.1002/adma.202106780}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-276403}, year = {2022}, abstract = {The extracellular matrix (ECM) of soft tissues in vivo has remarkable biological and structural properties. Thereby, the ECM provides mechanical stability while it still can be rearranged via cellular remodeling during tissue maturation or healing processes. However, modern synthetic alternatives fail to provide these key features among basic properties. Synthetic matrices are usually completely degraded or are inert regarding cellular remodeling. Based on a refined electrospinning process, a method is developed to generate synthetic scaffolds with highly porous fibrous structures and enhanced fiber-to-fiber distances. Since this approach allows for cell migration, matrix remodeling, and ECM synthesis, the scaffold provides an ideal platform for the generation of soft tissue equivalents. Using this matrix, an electrospun-based multilayered skin equivalent composed of a stratified epidermis, a dermal compartment, and a subcutis is able to be generated without the use of animal matrix components. The extension of classical dense electrospun scaffolds with high porosities and motile fibers generates a fully synthetic and defined alternative to collagen-gel-based tissue models and is a promising system for the construction of tissue equivalents as in vitro models or in vivo implants.}, language = {en} } @article{WussmannGroeberBeckerRiedletal.2022, author = {Wußmann, Maximiliane and Groeber-Becker, Florian Kai and Riedl, Sabrina and Alihodzic, Dina and Padaric, Daniel and Gerlitz, Lisa and Stallinger, Alexander and Liegl-Atzwanger, Bernadette and Zweytick, Dagmar and Rinner, Beate}, title = {In model, in vitro and in vivo killing efficacy of antitumor peptide RDP22 on MUG-Mel2, a patient derived cell line of an aggressive melanoma metastasis}, series = {Biomedicines}, volume = {10}, journal = {Biomedicines}, number = {11}, issn = {2227-9059}, doi = {10.3390/biomedicines10112961}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-297525}, year = {2022}, abstract = {The host defense derived peptide was assessed in different model systems with increasing complexity employing the highly aggressive NRAS mutated melanoma metastases cell line MUG-Mel2. Amongst others, fluorescence microscopy and spectroscopy, as well as cell death studies were applied for liposomal, 2D and 3D in vitro models including tumor spheroids without or within skin models and in vivo mouse xenografts. Summarized, MUG-Mel2 cells were shown to significantly expose the negatively charged lipid phosphatidylserine on their plasma membranes, showing they are successfully targeted by RDP22. The peptide was able to induce cell death in MUG-Mel2 2D and 3D cultures, where it was able to kill tumor cells even inside the core of tumor spheroids or inside a melanoma organotypic model. In vitro studies indicated cell death by apoptosis upon peptide treatment with an LC\(_{50}\) of 8.5 µM and seven-fold specificity for the melanoma cell line MUG-Mel2 over normal dermal fibroblasts. In vivo studies in mice xenografts revealed effective tumor regression upon intratumoral peptide injection, indicated by the strong clearance of pigmented tumor cells and tremendous reduction in tumor size and proliferation, which was determined histologically. The peptide RDP22 has clearly shown high potential against the melanoma cell line MUG-Mel2 in vitro and in vivo.}, language = {en} } @article{ReuterHaufImdahletal.2023, author = {Reuter, Christian and Hauf, Laura and Imdahl, Fabian and Sen, Rituparno and Vafadarnejad, Ehsan and Fey, Philipp and Finger, Tamara and Jones, Nicola G. and Walles, Heike and Barquist, Lars and Saliba, Antoine-Emmanuel and Groeber-Becker, Florian and Engstler, Markus}, title = {Vector-borne Trypanosoma brucei parasites develop in artificial human skin and persist as skin tissue forms}, series = {Nature Communications}, volume = {14}, journal = {Nature Communications}, doi = {10.1038/s41467-023-43437-2}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-358142}, year = {2023}, abstract = {Transmission of Trypanosoma brucei by tsetse flies involves the deposition of the cell cycle-arrested metacyclic life cycle stage into mammalian skin at the site of the fly's bite. We introduce an advanced human skin equivalent and use tsetse flies to naturally infect the skin with trypanosomes. We detail the chronological order of the parasites' development in the skin by single-cell RNA sequencing and find a rapid activation of metacyclic trypanosomes and differentiation to proliferative parasites. Here we show that after the establishment of a proliferative population, the parasites enter a reversible quiescent state characterized by slow replication and a strongly reduced metabolism. We term these quiescent trypanosomes skin tissue forms, a parasite population that may play an important role in maintaining the infection over long time periods and in asymptomatic infected individuals.}, language = {en} }