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- ItemNon-leaching, Highly Biocompatible Nanocellulose Surfaces That Efficiently Resist Fouling by Bacteria in an Artificial Dermis Model(Washington, DC : ACS Publications, 2020) Hassan, Ghada; Forsman, Nina; Wan, Xing; Keurulainen, Leena; Bimbo, Luis M.; Stehl, Susanne; van Charante, Frits; Chrubasik, Michael; Prakash, Aruna S.; Johansson, Leena-Sisko; Mullen, Declan C.; Johnston, Blair F.; Zimmermann, Ralf; Werner, Carsten; Yli-Kauhaluoma, Jari; Coenye, Tom; Saris, Per E.J.; Österberg, Monika; Moreira, Vânia M.Bacterial biofilm infections incur massive costs on healthcare systems worldwide. Particularly worrisome are the infections associated with pressure ulcers and prosthetic, plastic, and reconstructive surgeries, where staphylococci are the major biofilm-forming pathogens. Non-leaching antimicrobial surfaces offer great promise for the design of bioactive coatings to be used in medical devices. However, the vast majority are cationic, which brings about undesirable toxicity. To circumvent this issue, we have developed antimicrobial nanocellulose films by direct functionalization of the surface with dehydroabietic acid derivatives. Our conceptually unique design generates non-leaching anionic surfaces that reduce the number of viable staphylococci in suspension, including drug-resistant Staphylococcus aureus, by an impressive 4-5 log units, upon contact. Moreover, the films clearly prevent bacterial colonization of the surface in a model mimicking the physiological environment in chronic wounds. Their activity is not hampered by high protein content, and they nurture fibroblast growth at the surface without causing significant hemolysis. In this work, we have generated nanocellulose films with indisputable antimicrobial activity demonstrated using state-of-the-art models that best depict an "in vivo scenario". Our approach is to use fully renewable polymers and find suitable alternatives to silver and cationic antimicrobials. © 2020 American Chemical Society.
- ItemBlend Structure and n-Type Thermoelectric Performance of PA6/SAN and PA6/PMMA Blends Filled with Singlewalled Carbon Nanotubes(Basel : MDPI, 2021-4-28) Krause, Beate; Liguoro, Alice; Pötschke, PetraThe present study investigates how the formation of melt-mixed immiscible blends based on PA6/SAN and PA6/PMMA filled with single walled nanotubes (SWCNTs) affects the thermoelectric (TE) properties. In addition to the detailed investigation of the blend morphology with compositions between 100/0 wt.% and 50/50 wt.%, the thermoelectric properties are investigated on blends with different SWCNT concentrations (0.25–3.0 wt.%). Both PA6 and the blend composites with the used type of SWCNTs showed negative Seebeck coefficients. It was shown that the PA6 matrix polymer, in which the SWCNTs are localized, mainly influenced the thermoelectric properties of blends with high SWCNT contents. By varying the blend composition, an increase in the absolute Seebeck coefficient, power factor (PF), and figure of merit (ZT) was achieved compared to the PA6 composite which is mainly related to the selective localization and enrichment of SWCNTs in the PA6 matrix at constant SWCNT loading. The maximum PFs achieved were 0.22 µW/m·K2 for PA6/SAN/SWCNT 70/30/3 wt.% and 0.13 µW/m·K2 for PA6/PMMA/SWCNT 60/40/3 wt.% compared to 0.09 µW/m·K2 for PA6/3 wt.% SWCNT which represent increases to 244% and 144%, respectively. At higher PMMA or SAN concentration, the change from matrix-droplet to a co-continuous morphology started, which, despite higher SWCNT enrichment in the PA6 matrix, disturbed the electrical conductivity, resulting in reduced PFs with still increasing Seebeck coefficients. At SWCNT contents between 0.5 and 3 wt.% the increase in the absolute Seebeck coefficient was compensated by lower electrical conductivity resulting in lower PF and ZT as compared to the PA6 composites.
- ItemSurface-Dependent Osteoblasts Response to TiO2 Nanotubes of Dfferent Crystallinity(Basel : MDPI, 2020) Khrunyk, Yuliya Y.; Belikov, Sergey V.; Tsurkan, Mikhail V.; Vyalykh, Ivan V.; Markaryan, Alexandr Y.; Karabanalov, Maxim S.; Popov, Artemii A.; Wysokowski, MarcinOne of the major challenges of implantology is to design nanoscale modifications of titanium implant surfaces inducing osseointegration. The aim of this study was to investigate the behavior of rat osteoblasts cultured on anodized TiO2 nanotubes of different crystallinity (amorphous and anatase phase) up to 24 days. TiO2 nanotubes were fabricated on VT1–0 titanium foil via a two-step anodization at 20 V using NH4F as an electrolyte. Anatase-phase samples were prepared by heat treatment at 500 °C for 1 h. VT1–0 samples with flat surfaces were used as controls. Primary rat osteoblasts were seeded over experimental surfaces for several incubation times. Scanning electron microscopy (SEM) was used to analyze tested surfaces and cell morphology. Cell adhesion and proliferation were investigated by cell counting. Osteogenic differentiation of cells was evaluated by qPCR of runt-related transcription factor 2 (RUNX2), osteopontin (OPN), integrin binding sialoprotein (IBSP), alkaline phosphatase (ALP) and osteocalcin (OCN). Cell adhesion and proliferation, cell morphology and the expression of osteogenic markers were affected by TiO2 nanotube layered substrates of amorphous and anatase crystallinity. In comparison with flat titanium, along with increased cell adhesion and cell growth a large portion of osteoblasts grown on the both nanostructured surfaces exhibited an osteocyte-like morphology as early as 48 h of culture. Moreover, the expression of all tested osteogenic markers in cells cultured on amorphous and anatase TiO2 nanotubes was upregulated at least at one of the analyzed time points. To summarize, we demonstrated that amorphous and anodized TiO2 layered substrates are highly biocompatible with rat osteoblasts and that the surface modification with about 1500 nm length nanotubes of 35 ± 4 (amorphous phase) and 41 ± 8 nm (anatase phase) in diameter is sufficient to induce their osteogenic differentiation. Such results are significant to the engineering of coating strategies for orthopedic implants aimed to establish a more efficient bone to implant contact and enhance bone repair. © 2020 by the author. Licensee MDPI, Basel, Switzerland.
- ItemMaintenance of Ligament Homeostasis of Spheroid-Colonized Embroidered and Functionalized Scaffolds after 3D Stretch(Basel : Molecular Diversity Preservation International (MDPI), 2021) Gögele, Clemens; Konrad, Jens; Hahn, Judith; Breier, Annette; Schröpfer, Michaela; Meyer, Michael; Merkel, Rudolf; Hoffmann, Bernd; Schulze-Tanzil, GundulaAnterior cruciate ligament (ACL) ruptures are usually treated with autograft implantation to prevent knee instability. Tissue engineered ACL reconstruction is becoming promising to circumvent autograft limitations. The aim was to evaluate the influence of cyclic stretch on lapine (L) ACL fibroblasts on embroidered scaffolds with respect to adhesion, DNA and sulphated glycosaminoglycan (sGAG) contents, gene expression of ligament-associated extracellular matrix genes, such as type I collagen, decorin, tenascin C, tenomodulin, gap junctional connexin 43 and the transcription factor Mohawk. Control scaffolds and those functionalized by gas phase fluorination and cross-linked collagen foam were either pre-cultured with a suspension or with spheroids of LACL cells before being subjected to cyclic stretch (4%, 0.11 Hz, 3 days). Stretch increased significantly the scaffold area colonized with cells but impaired sGAGs and decorin gene expression (functionalized scaffolds seeded with cell suspension). Stretching increased tenascin C, connexin 43 and Mohawk but decreased decorin gene expression (control scaffolds seeded with cell suspension). Pre-cultivation of functionalized scaffolds with spheroids might be the more suitable method for maintaining ligamentogenesis in 3D scaffolds compared to using a cell suspension due to a significantly higher sGAG content in response to stretching and type I collagen gene expression in functionalized scaffolds.
- ItemPoly(lysine) Dendrimers Form Complexes with siRNA and Provide Its Effcient Uptake by Myeloid Cells: Model Studies for Therapeutic Nucleic Acid Delivery(Basel : Molecular Diversity Preservation International, 2020) Gorzkiewicz, Michał; Kopeć, Olga; Janaszewska, Anna; Konopka, Małgorzata; Pędziwiatr-Werbicka, Elżbieta; Tarasenko, Irina I.; Bezrodnyi, Valeriy V.; Neelov, Igor M.; Klajnert-Maculewicz, BarbaraThe disruption of the cellular pathways of protein biosynthesis through the mechanism of RNA interference has been recognized as a tool of great diagnostic and therapeutic significance. However, in order to fully exploit the potential of this phenomenon, efficient and safe carriers capable of overcoming extra-and intracellular barriers and delivering siRNA to the target cells are needed. Recently, attention has focused on the possibility of the application of multifunctional nanoparticles, dendrimers, as potential delivery devices for siRNA. The aim of the present work was to evaluate the formation of dendriplexes using novel poly(lysine) dendrimers (containing lysine and arginine or histidine residues in their structure), and to verify the hypothesis that the use of these polymers may allow an efficient method of siRNA transfer into the cells in vitro to be obtained. The fluorescence polarization studies, as well as zeta potential and hydrodynamic diameter measurements were used to characterize the dendrimer:siRNA complexes. The cytotoxicity of dendrimers and dendriplexes was evaluated with the resazurin-based assay. Using the flow cytometry technique, the efficiency of siRNA transport to the myeloid cells was determined. This approach allowed us to determine the properties and optimal molar ratios of dendrimer:siRNA complexes, as well as to demonstrate that poly(lysine) dendrimers may serve as efficient carriers of genetic material, being much more effective than the commercially available transfection agent Lipofectamine 2000. This outcome provides the basis for further research on the application of poly(lysine) dendrimers as carriers for nucleic acids in the field of gene therapy. © 2020 by the authors.
- ItemPreclinical Testing of New Hydrogel Materials for Cartilage Repair: Overcoming Fixation Issues in a Large Animal Model(New York, NY [u.a.] : Hindawi Publ. Corp., 2021) Lotz, Benedict; Bothe, Friederike; Deubel, Anne-Kathrin; Hesse, Eliane; Renz, Yvonne; Werner, Carsten; Schäfer, Simone; Böck, Thomas; Groll, Jürgen; von Rechenberg, Brigitte; Richter, Wiltrud; Hagmann, SebastienReinforced hydrogels represent a promising strategy for tissue engineering of articular cartilage. They can recreate mechanical and biological characteristics of native articular cartilage and promote cartilage regeneration in combination with mesenchymal stromal cells. One of the limitations of in vivo models for testing the outcome of tissue engineering approaches is implant fixation. The high mechanical stress within the knee joint, as well as the concave and convex cartilage surfaces, makes fixation of reinforced hydrogel challenging. Methods. Different fixation methods for full-thickness chondral defects in minipigs such as fibrin glue, BioGlue®, covering, and direct suturing of nonenforced and enforced constructs were compared. Because of insufficient fixation in chondral defects, superficial osteochondral defects in the femoral trochlea, as well as the femoral condyle, were examined using press-fit fixation. Two different hydrogels (starPEG and PAGE) were compared by 3D-micro-CT (μCT) analysis as well as histological analysis. Results. Our results showed fixation of below 50% for all methods in chondral defects. A superficial osteochondral defect of 1 mm depth was necessary for long-term fixation of a polycaprolactone (PCL)-reinforced hydrogel construct. Press-fit fixation seems to be adapted for a reliable fixation of 95% without confounding effects of glue or suture material. Despite the good integration of our constructs, especially in the starPEG group, visible bone lysis was detected in micro-CT analysis. There was no significant difference between the two hydrogels (starPEG and PAGE) and empty control defects regarding regeneration tissue and cell integration. However, in the starPEG group, more cell-containing hydrogel fragments were found within the defect area. Conclusion. Press-fit fixation in a superficial osteochondral defect in the medial trochlear groove of adult minipigs is a promising fixation method for reinforced hydrogels. To avoid bone lysis, future approaches should focus on multilayered constructs recreating the zonal cartilage as well as the calcified cartilage and the subchondral bone plate.
- ItemEnhanced growth of lapine anterior cruciate ligament-derived fibroblasts on scaffolds embroidered from poly(L-lactide-co-ε-caprolactone) and polylactic acid threads functionalized by fluorination and hexamethylene diisocyanate cross-linked collagen foams(Basel : Molecular Diversity Preservation International, 2020) Gögele, Clemens; Hahn, Judith; Elschner, Cindy; Breier, Annette; Schröpfer, Michaela; Prade, Ina; Meyer, Michael; Schulze-Tanzil, GundulaReconstruction of ruptured anterior cruciate ligaments (ACLs) is limited by the availability and donor site morbidity of autografts. Hence, a tissue engineered graft could present an alternative in the future. This study was undertaken to determine the performance of lapine (L) ACL-derived fibroblasts on embroidered poly(l-lactide-co-e-caprolactone) (P(LA-CL)) and polylactic acid (PLA) scaffolds in regard to a tissue engineering approach for ACL reconstruction. Surface modifications of P(LA-CL)/PLA by gas-phase fluorination and cross-linking of a collagen foam using either ethylcarbodiimide (EDC) or hexamethylene diisocyanate (HMDI) were tested regarding their influence on cell adhesion, growth and gene expression. The experiments were performed using embroidered P(LA-CL)/PLA scaffolds that were seeded dynamically or statically with LACL-derived fibroblasts. Scaffold cytocompatibility, cell survival, numbers, metabolic activity, ultrastructure and sulfated glycosaminoglycan (sGAG) synthesis were evaluated. Quantitative real-time polymerase chain reaction (QPCR) revealed gene expression of collagen type I (COL1A1), decorin (DCN), tenascin C (TNC), Mohawk (MKX) and tenomodulin (TNMD). All tested scaffolds were highly cytocompatible. A significantly higher cellularity and larger scaffold surface areas colonized by cells were detected in HMDI cross-linked and fluorinated scaffolds compared to those cross-linked with EDC or without any functionalization. By contrast, sGAG synthesis was higher in controls. Despite the fact that the significance level was not reached, gene expressions of ligament extracellular matrix components and differentiation markers were generally higher in fluorinated scaffolds with cross-linked collagen foams. LACL-derived fibroblasts maintained their differentiated phenotype on fluorinated scaffolds supplemented with a HMDI cross-linked collagen foam, making them a promising tool for ACL tissue engineering. © 2020 by the authors. Licensee MDPI, Basel, Switzerland.
- ItemIn search of a phosphorus dendrimer-based carrier of rose bengal: Tyramine linker limits fluorescent and phototoxic properties of a photosensitizer(Basel : Molecular Diversity Preservation International, 2020) Sztandera, Krzysztof; Marcinkowska, Monika; Gorzkiewicz, Michał; Janaszewska, Anna; Laurent, Regis; Zabłocka, Maria; Mignani, Serge; Majoral, Jean Pierre; Klajnert-Maculewicz, BarbaraPhotodynamic therapy (PDT) is a skin cancer treatment alternative to chemotherapy and radiotherapy. This method exploits three elements: a phototoxic compound (photosensitizer), light source and oxygen. Upon irradiation by light of a specific wavelength, the photosensitizer generates reactive oxygen species triggering the cascade of reactions leading to cell death. The positive therapeutic effect of PDT may be limited due to low solubility, low tumor specificity and inefficient cellular uptake of photosensitizers. A promising approach to overcome these obstacles involves the use of nanocarrier systems. The aim of this initial study was to determine the potential of the application of phosphorus dendrimers as carriers of a photosensitizer—rose bengal (RB). The primary goal involved the synthesis and in vitro studies of covalent drug–dendrimer conjugates. Our approach allowed us to obtain RB–dendrimer conjugates with the use of tyramine as an aromatic linker between the carrier and the drug. The compounds were characterized by FT-IR,1H NMR,13C NMR,31P NMR, size and zeta potential measurements and spectrofluorimetric analysis. The dialysis to check the drug release from the conjugate, flow cytometry to specify intracellular uptake, and singlet oxygen generation assay were also applied. Finally, we used MTT assay to determine the biological activity of the tested compounds. The results of our experiments indicate that the conjugation of RB to phosphorus dendrimers via the tyramine linker decreases photodynamic activity of RB. © 2020 by the authors. Licensee MDPI, Basel, Switzerland.
- ItemSv40 transfected human anterior cruciate ligament derived ligamentocytes—suitable as a human in vitro model for ligament reconstruction?(Basel : Molecular Diversity Preservation International, 2020) Schulze-Tanzil, Gundula; Arnold, Philipp; Gögele, Clemens; Hahn, Judith; Breier, Annette; Meyer, Michael; Kohl, Benjamin; Schröpfer, Michaela; Schwarz, SilkeCultured human primary cells have a limited lifespan undergoing dedifferentiation or senescence. Anterior cruciate ligaments (ACL) are hypocellular but tissue engineering (TE) requires high cell numbers. Simian virus (SV) 40 tumor (T) antigen expression could extend the lifespan of cells. This study aimed to identify cellular changes induced by SV40 expression in human ACL ligamentocytes by comparing them with non-transfected ligamentocytes and tissue of the same donor to assess their applicability as TE model. Human ACL ligamentocytes (40-year-old female donor after ACL rupture) were either transfected with a SV40 plasmid or remained non-transfected (control) before monitored for SV40 expression, survival, and DNA content. Protein expression of cultured ligamentocytes was compared with the donor tissue. Ligamentocyte spheroids were seeded on scaffolds embroidered either from polylactic acid (PLA) threads solely or combined PLA and poly (L-lactide-co-e-caprolactone) (P(LA-CL)) threads. These scaffolds were further functionalized with fluorination and fibrillated collagen foam. Cell distribution and survival were monitored for up to five weeks. The transfected cells expressed the SV40 antigen throughout the entire observation time, but often exhibited random and incomplete cell divisions with significantly more dying cells, significantly more DNA and more numerous nucleoli than controls. The expression profile of non-transfected and SV40-positive ligamentocytes was similar. In contrast to controls, SV40-positive cells formed larger spheroids, produced less vimentin and focal adhesions and died on the scaffolds after 21 d. Functionalized scaffolds supported human ligamentocyte growth. SV40 antigen expressing ligamentocytes share many properties with their non-transfected counterparts suggesting them as a model, however, applicability for TE is limited. © 2020 by the authors. Licensee MDPI, Basel, Switzerland.
- ItemTuning the Interactions in Multiresponsive Complex Coacervate-Based Underwater Adhesives(Basel : Molecular Diversity Preservation International, 2020) Dompé, Marco; Cedano-Serrano, Francisco J.; Vahdati, Mehdi; Sidoli, Ugo; Heckert, Olaf; Synytska, Alla; Hourdet, Dominique; Creton, Costantino; van der Gucht, Jasper; Kodger, Thomas; Kamperman, MarleenIn this work, we report the systematic investigation of a multiresponsive complex coacervate-based underwater adhesive, obtained by combining polyelectrolyte domains and thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) units. This material exhibits a transition from liquid to solid but, differently from most reactive glues, is completely held together by non-covalent interactions, i.e., electrostatic and hydrophobic. Because the solidification results in a kinetically trapped morphology, the final mechanical properties strongly depend on the preparation conditions and on the surrounding environment. A systematic study is performed to assess the effect of ionic strength and of PNIPAM content on the thermal, rheological and adhesive properties. This study enables the optimization of polymer composition and environmental conditions for this underwater adhesive system. The best performance with a work of adhesion of 6.5 J/m2 was found for the complex coacervates prepared at high ionic strength (0.75 M NaCl) and at an optimal PNIPAM content around 30% mol/mol. The high ionic strength enables injectability, while the hydrated PNIPAM domains provide additional dissipation, without softening the material so much that it becomes too weak to resist detaching stress. © 2019 by the authors. Licensee MDPI, Basel, Switzerland.