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- ItemIn vivo ligamentogenesis in embroidered poly(lactic-co-ε-caprolactone) / polylactic acid scaffolds functionalized by fluorination and hexamethylene diisocyanate cross-linked collagen foams(Berlin ; Heidelberg : Springer, 2022) Kokozidou, Maria; Gögele, Clemens; Pirrung, Felix; Hammer, Niels; Werner, Christian; Kohl, Benjamin; Hahn, Judith; Breier, Annette; Schröpfer, Michaela; Meyer, Michael; Schulze-Tanzil, GundulaAlthough autografts represent the gold standard for anterior cruciate ligament (ACL) reconstruction, tissue-engineered ACLs provide a prospect to minimize donor site morbidity and limited graft availability. This study characterizes the ligamentogenesis in embroidered poly(L-lactide-co-epsilon-caprolactone) (P(LA-CL)) / polylactic acid (PLA) constructs using a dynamic nude mice xenograft model. (P(LA-CL))/PLA scaffolds remained either untreated (co) or were functionalized by gas fluorination (F), collagen foam cross-linked with hexamethylene diisocyanate (HMDI) (coll), or F combined with the foam (F+coll). Cell-free constructs or those seeded for 1 week with lapine ACL ligamentocytes were implanted into nude mice for 12 weeks. Following explantation, cell vitality and content, histo(patho)logy of scaffolds (including organs: liver, kidney, spleen), sulphated glycosaminoglycan (sGAG) contents and biomechanical properties were assessed. Scaffolds did not affect mice weight development and organs, indicating no organ toxicity. Moreover, scaffolds maintained their size and shape and reflected a high cell viability prior to and following implantation. Coll or F+coll scaffolds seeded with cells yielded superior macroscopic properties compared to the controls. Mild signs of inflammation (foreign-body giant cells and hyperemia) were limited to scaffolds without collagen. Microscopical score values and sGAG content did not differ significantly. Although remaining stable after explantation, elastic modulus, maximum force, tensile strength and strain at F-max were significantly lower in explanted scaffolds compared to those before implantation, with no significant differences between scaffold subtypes, except for a higher maximum force in F+coll compared with F samples (in vivo). Scaffold functionalization with fluorinated collagen foam provides a promising approach for ACL tissue engineering.
- ItemPseudo-chemotaxis of active Brownian particles competing for food(San Francisco, California, US : PLOS, 2020) Merlitz, Holger; Vuijk, Hidde D.; Wittmann, René; Sharma, Abhinav; Sommer, Jens-UweActive Brownian particles (ABPs) are physical models for motility in simple life forms and easily studied in simulations. An open question is to what extent an increase of activity by a gradient of fuel, or food in living systems, results in an evolutionary advantage of actively moving systems such as ABPs over non-motile systems, which rely on thermal diffusion only. It is an established fact that within confined systems in a stationary state, the activity of ABPs generates density profiles that are enhanced in regions of low activity, which is thus referred to as ‘anti-chemotaxis’. This would suggest that a rather complex sensoric subsystem and information processing is a precondition to recognize and navigate towards a food source. We demonstrate in this work that in non-stationary setups, for instance as a result of short bursts of fuel/food, ABPs do in fact exhibit chemotactic behavior. In direct competition with inactive, but otherwise identical Brownian particles (BPs), the ABPs are shown to fetch a larger amount of food. We discuss this result based on simple physical arguments. From the biological perspective, the ability of primitive entities to move in direct response to the available amount of external energy would, even in absence of any sensoric devices, encompass an evolutionary advantage.
- ItemStructure, Properties, and Release Kinetics of the Polymer/Insect Repellent System Poly (l-Lactic Acid)/Ethyl Butylacetylaminopropionate (PLLA/IR3535)(Basel : MDPI, 2022) Du, Fanfan; Erdmann, Rafael; Petzold, Albrecht; Wutzler, Andre; Leuteritz, Andreas; Nase, Michael; Androsch, RenéThe insect repellent ethyl butylacetylaminopropionate (IR3535) was used as a functional additive for poly (l-lactic acid) (PLLA) to modify its structure and mechanical properties and achieve insect repellency. PLLA/IR3535 mixtures at various compositions were prepared via melt extrusion. In the analyzed composition range of 0 to 23 m% IR3535, PLLA and IR3535 were miscible at the length scale represented by the glass transition temperature. Addition of IR3535 resulted in a significant decrease in the glass transition temperature of PLLA, as well as in the elastic modulus, indicating its efficiency as a plasticizer. All mixtures were amorphous after extrusion, though PLLA/IR3535 extrudates with an IR3535 content between 18 and 23 m% crystallized during long-term storage at ambient temperature, due to their low glass transition temperature. Quantification of the release of IR3535 into the environment by thermogravimetric analysis at different temperatures between 50 and 100 °C allowed the estimation of the evaporation rate at lower temperatures, suggesting an extremely low release rate with a time constant of the order of magnitude of 1–2 years at body temperature.
- ItemTargeted RNAi of BIRC5/Survivin Using Antibody-Conjugated Poly(Propylene Imine)-Based Polyplexes Inhibits Growth of PSCA-Positive Tumors(Basel : MDPI, 2021) Jugel, Willi; Aigner, Achim; Michen, Susanne; Hagstotz, Alexander; Ewe, Alexander; Appelhans, Dietmar; Schackert, Gabriele; Temme, Achim; Tietze, StefanieDelivery of siRNAs for the treatment of tumors critically depends on the development of efficient nucleic acid carrier systems. The complexation of dendritic polymers (dendrimers) results in nanoparticles, called dendriplexes, that protect siRNA from degradation and mediate non-specific cellular uptake of siRNA. However, large siRNA doses are required for in vivo use due to accumulation of the nanoparticles in sinks such as the lung, liver, and spleen. This suggests the exploration of targeted nanoparticles for enhancing tumor cell specificity and achieving higher siRNA levels in tumors. In this work, we report on the targeted delivery of a therapeutic siRNA specific for BIRC5/Survivin in vitro and in vivo to tumor cells expressing the surface marker prostate stem cell antigen (PSCA). For this, polyplexes consisting of single-chain antibody fragments specific for PSCA conjugated to siRNA/maltose-modified poly(propylene imine) dendriplexes were used. These polyplexes were endocytosed by PSCA-positive 293TPSCA/ffLuc and PC3PSCA cells and caused knockdown of reporter gene firefly luciferase and Survivin expression, respectively. In a therapeutic study in PC3PSCA xenograft-bearing mice, significant anti-tumor effects were observed upon systemic administration of the targeted polyplexes. This indicates superior anti-tumor efficacy when employing targeted delivery of Survivin-specific siRNA, based on the additive effects of siRNA-mediated Survivin knockdown in combination with scFv-mediated PSCA inhibition.
- ItemA Complementary and Revised View on the N-Acylation of Chitosan with Hexanoyl Chloride(Basel : MDPI, 2021) Reis, Berthold; Gerlach, Niklas; Steinbach, Christine; Haro Carrasco, Karina; Oelmann, Marina; Schwarz, Simona; Müller, Martin; Schwarz, DanaThe modification of the biobased polymer chitosan is a broad and widely studied field. Herein, an insight into the hydrophobization of low-molecular-weight chitosan by substitution of amino functionalities with hexanoyl chloride is reported. Thereby, the influence of the pH of the reaction media was investigated. Further, methods for the determination of the degree of substitution based on 1H-NMR, FTIR, and potentiometric titration were compared and discussed regarding their accuracy and precision. 1H-NMR was the most accurate method, while FTIR and the potentiometric titration, though precise and reproducible, underlie the influence of complete protonation and solubility issues. Additionally, the impact of the pH variation during the synthesis on the properties of the samples was investigated by Cd2+ sorption experiments. The adjusted pH values during the synthesis and, therefore, the obtained degrees of substitution possessed a strong impact on the adsorption properties of the final material.
- ItemElectrochemical approach for isolation of chitin from the skeleton of the black coral cirrhipathes sp. (Antipatharia)(Basel : MDPI, 2020) Nowacki, Krzysztof; Stępniak, Izabela; Langer, Enrico; Tsurkan, Mikhail; Wysokowski, Marcin; Petrenko, Iaroslav; Khrunyk, Yuliya; Fursov, Andriy; Bo, Marzia; Bavestrello, Giorgio; Joseph, Yvonne; Ehrlich, HermannThe development of novel and effective methods for the isolation of chitin, which remains one of the fundamental aminopolysaccharides within skeletal structures of diverse marine invertebrates, is still relevant. In contrast to numerous studies on chitin extraction from crustaceans, mollusks and sponges, there are only a few reports concerning its isolation from corals, and especially black corals (Antipatharia). In this work, we report the stepwise isolation and identification of chitin from Cirrhipathes sp. (Antipatharia, Antipathidae) for the first time. The proposed method, aiming at the extraction of the chitinous scaffold from the skeleton of black coral species, combined a well-known chemical treatment with in situ electrolysis, using a concentrated Na2SO4 aqueous solution as the electrolyte. This novel method allows the isolation of a-chitin in the form of a microporous membrane-like material. Moreover, the extracted chitinous scaffold, with a well-preserved, unique pore distribution, has been extracted in an astoundingly short time (12 h) compared to the earlier reported attempts at chitin isolation from Antipatharia corals. © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
- ItemPoly(2-alkyl-2-oxazoline)-Heparin Hydrogels—Expanding the Physicochemical Parameter Space of Biohybrid Materials(Weinheim : Wiley-VCH, 2021) Hahn, Dominik; Sonntag, Jannick M.; Lück, Steffen; Maitz, Manfred F.; Freudenberg, Uwe; Jordan, Rainer; Werner, CarstenPoly(ethylene glycol) (PEG)-glycosaminoglycan (GAG) hydrogel networks are established as very versatile biomaterials. Herein, the synthetic gel component of the biohybrid materials is systematically varied by combining different poly(2-alkyl-2-oxazolines) (POx) with heparin applying a Michael-type addition crosslinking scheme: POx of gradated hydrophilicity and temperature-responsiveness provides polymer networks of distinctly different stiffness and swelling. Adjusting the mechanical properties and the GAG concentration of the gels to similar values allows for modulating the release of GAG-binding growth factors (VEGF165 and PDGF-BB) by the choice of the POx and its temperature-dependent conformation. Adsorption of fibronectin, growth of fibroblasts, and bacterial adhesion scale with the hydrophobicity of the gel-incorporated POx. In vitro hemocompatibility tests with freshly drawn human whole blood show advantages of POx-based gels compared to the PEG-based reference materials. Biohybrid POx hydrogels can therefore enable biomedical technologies requiring GAG-based materials with customized and switchable physicochemical characteristics. © 2021 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.
- ItemBone marrow mesenchymal stromal cell-derived extracellular matrix displays altered glycosaminoglycan structure and impaired functionality in Myelodysplastic Syndromes(Lausanne : Frontiers Media, 2022) Bains, Amanpreet Kaur; Behrens Wu, Lena; Rivière, Jennifer; Rother, Sandra; Magno, Valentina; Friedrichs, Jens; Werner, Carsten; Bornhäuser, Martin; Götze, Katharina S.; Cross, Michael; Platzbecker, Uwe; Wobus, ManjaMyelodysplastic syndromes (MDS) comprise a heterogeneous group of hematologic malignancies characterized by clonal hematopoiesis, one or more cytopenias such as anemia, neutropenia, or thrombocytopenia, abnormal cellular maturation, and a high risk of progression to acute myeloid leukemia. The bone marrow microenvironment (BMME) in general and mesenchymal stromal cells (MSCs) in particular contribute to both the initiation and progression of MDS. However, little is known about the role of MSC-derived extracellular matrix (ECM) in this context. Therefore, we performed a comparative analysis of in vitro deposited MSC-derived ECM of different MDS subtypes and healthy controls. Atomic force microscopy analyses demonstrated that MDS ECM was significantly thicker and more compliant than those from healthy MSCs. Scanning electron microscopy showed a dense meshwork of fibrillar bundles connected by numerous smaller structures that span the distance between fibers in MDS ECM. Glycosaminoglycan (GAG) structures were detectable at high abundance in MDS ECM as white, sponge-like arrays on top of the fibrillar network. Quantification by Blyscan assay confirmed these observations, with higher concentrations of sulfated GAGs in MDS ECM. Fluorescent lectin staining with wheat germ agglutinin and peanut agglutinin demonstrated increased deposition of N-acetyl-glucosamine GAGs (hyaluronan (HA) and heparan sulfate) in low risk (LR) MDS ECM. Differential expression of N-acetyl-galactosamine GAGs (chondroitin sulfate, dermatan sulfate) was observed between LR- and high risk (HR)-MDS. Moreover, increased amounts of HA in the matrix of MSCs from LR-MDS patients were found to correlate with enhanced HA synthase 1 mRNA expression in these cells. Stimulation of mononuclear cells from healthy donors with low molecular weight HA resulted in an increased expression of various pro-inflammatory cytokines suggesting a contribution of the ECM to the inflammatory BMME typical of LR-MDS. CD34+ hematopoietic stem and progenitor cells (HSPCs) displayed an impaired differentiation potential after cultivation on MDS ECM and modified morphology accompanied by decreased integrin expression which mediate cell-matrix interaction. In summary, we provide evidence for structural alterations of the MSC-derived ECM in both LR- and HR-MDS. GAGs may play an important role in this remodeling processes during the malignant transformation which leads to the observed disturbance in the support of normal hematopoiesis.
- ItemReal-Time Monitoring of Blood Parameters in the Intensive Care Unit: State-of-the-Art and Perspectives(Basel : MDPI, 2022) Bockholt, Rebecca; Paschke, Shaleen; Heubner, Lars; Ibarlucea, Bergoi; Laupp, Alexander; Janićijević, Željko; Klinghammer, Stephanie; Balakin, Sascha; Maitz, Manfred F.; Werner, Carsten; Cuniberti, Gianaurelio; Baraban, Larysa; Spieth, Peter MarkusThe number of patients in intensive care units has increased over the past years. Critically ill patients are treated with a real time support of the instruments that offer monitoring of relevant blood parameters. These parameters include blood gases, lactate, and glucose, as well as pH and tem-perature. Considering the COVID-19 pandemic, continuous management of dynamic deteriorating parameters in patients is more relevant than ever before. This narrative review aims to summarize the currently available literature regarding real-time monitoring of blood parameters in intensive care. Both, invasive and non-invasive methods are described in detail and discussed in terms of general advantages and disadvantages particularly in context of their use in different medical fields but especially in critical care. The objective is to explicate both, well-known and frequently used as well as relatively unknown devices. Furtehrmore, potential future direction in research and development of realtime sensor systems are discussed. Therefore, the discussion section provides a brief description of current developments in biosensing with special emphasis on their technical implementation. In connection with these developments, the authors focus on different electrochemical approaches to invasive and non-invasive measurements in vivo.
- ItemA Practical Guide to the Automated Analysis of Vascular Growth, Maturation and Injury in the Brain(Lausanne : Frontiers Media, 2020) Rust, Ruslan; Kirabali, Tunahan; Grönnert, Lisa; Dogancay, Berre; Limasale, Yanuar D.P.; Meinhardt, Andrea; Werner, Carsten; Laviña, Bà rbara; Kulic, Luka; Nitsch, Roger M.; Tackenberg, Christian; Schwab, Martin E.The distinct organization of the brain’s vasculature ensures the adequate delivery of oxygen and nutrients during development and adulthood. Acute and chronic pathological changes of the vascular system have been implicated in many neurological disorders including stroke and dementia. Here, we describe a fast, automated method that allows the highly reproducible, quantitative assessment of distinct vascular parameters and their changes based on the open source software Fiji (ImageJ). In particular, we developed a practical guide to reliably measure aspects of growth, repair and maturation of the brain’s vasculature during development and neurovascular disease in mice and humans. The script can be used to assess the effects of different external factors including pharmacological treatments or disease states. Moreover, the procedure is expandable to blood vessels of other organs and vascular in vitro models. © Copyright © 2020 Rust, Kirabali, Grönnert, Dogancay, Limasale, Meinhardt, Werner, Laviña, Kulic, Nitsch, Tackenberg and Schwab.