@article{BorovaTokarevStahlhutetal.2020, author = {Borova, Solomiia and Tokarev, Victor and Stahlhut, Philipp and Luxenhofer, Robert}, title = {Crosslinking of hydrophilic polymers using polyperoxides}, series = {Colloid and Polymer Science}, volume = {298}, journal = {Colloid and Polymer Science}, doi = {10.1007/s00396-020-04738-w}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-238109}, pages = {1699-1713}, year = {2020}, abstract = {Hydrogels that can mimic mechanical properties and functions of biological tissue have attracted great interest in tissue engineering and biofabrication. In these fields, new materials and approaches to prepare hydrogels without using toxic starting materials or materials that decompose into toxic compounds remain to be sought after. Here, we report the crosslinking of commercial, unfunctionalized hydrophilic poly(2-ethyl-2-oxazoline) using peroxide copolymers in their melt. The influence of temperature, peroxide copolymer concentration, and duration of the crosslinking process has been investigated. The method allows to create hydrogels from unfunctionalized polymers in their melt and to control the mechanical properties of the resulting materials. The design of hydrogels with a suitable mechanical performance is of crucial importance in many existing and potential applications of soft materials, including medical applications.}, language = {en} } @article{HahnBeudertGutmannetal.2021, author = {Hahn, Lukas and Beudert, Matthias and Gutmann, Marcus and Keßler, Larissa and Stahlhut, Philipp and Fischer, Lena and Karakaya, Emine and Lorson, Thomas and Thievessen, Ingo and Detsch, Rainer and L{\"u}hmann, Tessa and Luxenhofer, Robert}, title = {From Thermogelling Hydrogels toward Functional Bioinks: Controlled Modification and Cytocompatible Crosslinking}, series = {Macromolecular Bioscience}, volume = {21}, journal = {Macromolecular Bioscience}, number = {10}, doi = {10.1002/mabi.202100122}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-257542}, year = {2021}, abstract = {Hydrogels are key components in bioink formulations to ensure printability and stability in biofabrication. In this study, a well-known Diels-Alder two-step post-polymerization modification approach is introduced into thermogelling diblock copolymers, comprising poly(2-methyl-2-oxazoline) and thermoresponsive poly(2-n-propyl-2-oxazine). The diblock copolymers are partially hydrolyzed and subsequently modified by acid/amine coupling with furan and maleimide moieties. While the thermogelling and shear-thinning properties allow excellent printability, trigger-less cell-friendly Diels-Alder click-chemistry yields long-term shape-fidelity. The introduced platform enables easy incorporation of cell-binding moieties (RGD-peptide) for cellular interaction. The hydrogel is functionalized with RGD-peptides using thiol-maleimide chemistry and cell proliferation as well as morphology of fibroblasts seeded on top of the hydrogels confirm the cell adhesion facilitated by the peptides. Finally, bioink formulations are tested for biocompatibility by incorporating fibroblasts homogenously inside the polymer solution pre-printing. After the printing and crosslinking process good cytocompatibility is confirmed. The established bioink system combines a two-step approach by physical precursor gelation followed by an additional chemical stabilization, offering a broad versatility for further biomechanical adaptation or bioresponsive peptide modification.}, language = {en} } @article{HaiderAhmadYangetal.2021, author = {Haider, Malik Salman and Ahmad, Taufiq and Yang, Mengshi and Hu, Chen and Hahn, Lukas and Stahlhut, Philipp and Groll, J{\"u}rgen and Luxenhofer, Robert}, title = {Tuning the thermogelation and rheology of poly(2-oxazoline)/poly(2-oxazine)s based thermosensitive hydrogels for 3D bioprinting}, series = {Gels}, volume = {7}, journal = {Gels}, number = {3}, issn = {2310-2861}, doi = {10.3390/gels7030078}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-241781}, year = {2021}, abstract = {As one kind of "smart" material, thermogelling polymers find applications in biofabrication, drug delivery and regenerative medicine. In this work, we report a thermosensitive poly(2-oxazoline)/poly(2-oxazine) based diblock copolymer comprising thermosensitive/moderately hydrophobic poly(2-N-propyl-2-oxazine) (pPrOzi) and thermosensitive/moderately hydrophilic poly(2-ethyl-2-oxazoline) (pEtOx). Hydrogels were only formed when block length exceeded certain length (≈100 repeat units). The tube inversion and rheological tests showed that the material has then a reversible sol-gel transition above 25 wt.\% concentration. Rheological tests further revealed a gel strength around 3 kPa, high shear thinning property and rapid shear recovery after stress, which are highly desirable properties for extrusion based three-dimensional (3D) (bio) printing. Attributed to the rheology profile, well resolved printability and high stackability (with added laponite) was also possible. (Cryo) scanning electron microscopy exhibited a highly porous, interconnected, 3D network. The sol-state at lower temperatures (in ice bath) facilitated the homogeneous distribution of (fluorescently labelled) human adipose derived stem cells (hADSCs) in the hydrogel matrix. Post-printing live/dead assays revealed that the hADSCs encapsulated within the hydrogel remained viable (≈97\%). This thermoreversible and (bio) printable hydrogel demonstrated promising properties for use in tissue engineering applications.}, language = {en} } @article{DoryabTaskinStahlhutetal.2021, author = {Doryab, Ali and Taskin, Mehmet Berat and Stahlhut, Philipp and Schr{\"o}ppel, Andreas and Wagner, Darcy E. and Groll, J{\"u}rgen and Schmid, Otmar}, title = {A Biomimetic, Copolymeric Membrane for Cell-Stretch Experiments with Pulmonary Epithelial Cells at the Air-Liquid Interface}, series = {Advanced Functional Materials}, volume = {31}, journal = {Advanced Functional Materials}, number = {10}, doi = {10.1002/adfm.202004707}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-225645}, year = {2021}, abstract = {Chronic respiratory diseases are among the leading causes of death worldwide, but only symptomatic therapies are available for terminal illness. This in part reflects a lack of biomimetic in vitro models that can imitate the complex environment and physiology of the lung. Here, a copolymeric membrane consisting of poly(ε-)caprolactone and gelatin with tunable properties, resembling the main characteristics of the alveolar basement membrane is introduced. The thin bioinspired membrane (≤5 μm) is stretchable (up to 25\% linear strain) with appropriate surface wettability and porosity for culturing lung epithelial cells under air-liquid interface conditions. The unique biphasic concept of this membrane provides optimum characteristics for initial cell growth (phase I) and then switch to biomimetic properties for cyclic cell-stretch experiments (phase II). It is showed that physiologic cyclic mechanical stretch improves formation of F-actin cytoskeleton filaments and tight junctions while non-physiologic over-stretch induces cell apoptosis, activates inflammatory response (IL-8), and impairs epithelial barrier integrity. It is also demonstrated that cyclic physiologic stretch can enhance the cellular uptake of nanoparticles. Since this membrane offers considerable advantages over currently used membranes, it may lead the way to more biomimetic in vitro models of the lung for translation of in vitro response studies into clinical outcome.}, language = {en} } @article{HuHahnYangetal.2021, author = {Hu, Chen and Hahn, Lukas and Yang, Mengshi and Altmann, Alexander and Stahlhut, Philipp and Groll, J{\"u}rgen and Luxenhofer, Robert}, title = {Improving printability of a thermoresponsive hydrogel biomaterial ink by nanoclay addition}, series = {Journal of Materials Science}, volume = {56}, journal = {Journal of Materials Science}, issn = {0022-2461}, doi = {10.1007/s10853-020-05190-5}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-234894}, pages = {691-705}, year = {2021}, abstract = {As a promising biofabrication technology, extrusion-based bioprinting has gained significant attention in the last decade and major advances have been made in the development of bioinks. However, suitable synthetic and stimuli-responsive bioinks are underrepresented in this context. In this work, we described a hybrid system of nanoclay Laponite XLG and thermoresponsive block copolymer poly(2-methyl-2-oxazoline)-b-poly(2-n-propyl-2-oxazine) (PMeOx-b-PnPrOzi) as a novel biomaterial ink and discussed its critical properties relevant for extrusion-based bioprinting, including viscoelastic properties and printability. The hybrid hydrogel retains the thermogelling properties but is strengthened by the added clay (over 5 kPa of storage modulus and 240 Pa of yield stress). Importantly, the shear-thinning character is further enhanced, which, in combination with very rapid viscosity recovery (~ 1 s) and structure recovery (~ 10 s), is highly beneficial for extrusion-based 3D printing. Accordingly, various 3D patterns could be printed with markedly enhanced resolution and shape fidelity compared to the biomaterial ink without added clay.}, language = {en} } @article{DoryabTaskinStahlhutetal.2021, author = {Doryab, Ali and Taskin, Mehmet Berat and Stahlhut, Philipp and Schr{\"o}ppel, Andreas and Orak, Sezer and Voss, Carola and Ahluwalia, Arti and Rehberg, Markus and Hilgendorff, Anne and St{\"o}ger, Tobias and Groll, J{\"u}rgen and Schmid, Otmar}, title = {A Bioinspired in vitro Lung Model to Study Particokinetics of Nano-/Microparticles Under Cyclic Stretch and Air-Liquid Interface Conditions}, series = {Frontiers in Bioengineering and Biotechnology}, volume = {9}, journal = {Frontiers in Bioengineering and Biotechnology}, issn = {2296-4185}, doi = {10.3389/fbioe.2021.616830}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-223830}, year = {2021}, abstract = {Evolution has endowed the lung with exceptional design providing a large surface area for gas exchange area (ca. 100 m\(^{2}\)) in a relatively small tissue volume (ca. 6 L). This is possible due to a complex tissue architecture that has resulted in one of the most challenging organs to be recreated in the lab. The need for realistic and robust in vitro lung models becomes even more evident as causal therapies, especially for chronic respiratory diseases, are lacking. Here, we describe the Cyclic In VItro Cell-stretch (CIVIC) "breathing" lung bioreactor for pulmonary epithelial cells at the air-liquid interface (ALI) experiencing cyclic stretch while monitoring stretch-related parameters (amplitude, frequency, and membrane elastic modulus) under real-time conditions. The previously described biomimetic copolymeric BETA membrane (5 μm thick, bioactive, porous, and elastic) was attempted to be improved for even more biomimetic permeability, elasticity (elastic modulus and stretchability), and bioactivity by changing its chemical composition. This biphasic membrane supports both the initial formation of a tight monolayer of pulmonary epithelial cells (A549 and 16HBE14o\(^{-}\)) under submerged conditions and the subsequent cell-stretch experiments at the ALI without preconditioning of the membrane. The newly manufactured versions of the BETA membrane did not improve the characteristics of the previously determined optimum BETA membrane (9.35\% PCL and 6.34\% gelatin [w/v solvent]). Hence, the optimum BETA membrane was used to investigate quantitatively the role of physiologic cyclic mechanical stretch (10\% linear stretch; 0.33 Hz: light exercise conditions) on size-dependent cellular uptake and transepithelial transport of nanoparticles (100 nm) and microparticles (1,000 nm) for alveolar epithelial cells (A549) under ALI conditions. Our results show that physiologic stretch enhances cellular uptake of 100 nm nanoparticles across the epithelial cell barrier, but the barrier becomes permeable for both nano- and micron-sized particles (100 and 1,000 nm). This suggests that currently used static in vitro assays may underestimate cellular uptake and transbarrier transport of nanoparticles in the lung.}, language = {en} } @article{HauptsteinForsterNadernezhadetal.2022, author = {Hauptstein, Julia and Forster, Leonard and Nadernezhad, Ali and Horder, Hannes and Stahlhut, Philipp and Groll, J{\"u}rgen and Blunk, Torsten and Teßmar, J{\"o}rg}, title = {Bioink Platform Utilizing Dual-Stage Crosslinking of Hyaluronic Acid Tailored for Chondrogenic Differentiation of Mesenchymal Stromal Cells}, series = {Macromolecular Bioscience}, volume = {22}, journal = {Macromolecular Bioscience}, number = {2}, doi = {10.1002/mabi.202100331}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-257556}, pages = {2100331}, year = {2022}, abstract = {3D bioprinting often involves application of highly concentrated polymeric bioinks to enable fabrication of stable cell-hydrogel constructs, although poor cell survival, compromised stem cell differentiation, and an inhomogeneous distribution of newly produced extracellular matrix (ECM) are frequently observed. Therefore, this study presents a bioink platform using a new versatile dual-stage crosslinking approach based on thiolated hyaluronic acid (HA-SH), which not only provides stand-alone 3D printability but also facilitates effective chondrogenic differentiation of mesenchymal stromal cells. A range of HA-SH with different molecular weights is synthesized and crosslinked with acrylated (PEG-diacryl) and allylated (PEG-diallyl) polyethylene glycol in a two-step reaction scheme. The initial Michael addition is used to achieve ink printability, followed by UV-mediated thiol-ene reaction to stabilize the printed bioink for long-term cell culture. Bioinks with high molecular weight HA-SH (>200 kDa) require comparably low polymer content to facilitate bioprinting. This leads to superior quality of cartilaginous constructs which possess a coherent ECM and a strongly increased stiffness of long-term cultured constructs. The dual-stage system may serve as an example to design platforms using two independent crosslinking reactions at one functional group, which allows adjusting printability as well as material and biological properties of bioinks.}, language = {en} } @article{BrandForsterBoecketal.2022, author = {Brand, Jessica S. and Forster, Leonard and B{\"o}ck, Thomas and Stahlhut, Philipp and Teßmar, J{\"o}rg and Groll, J{\"u}rgen and Albrecht, Krystyna}, title = {Covalently Cross-Linked Pig Gastric Mucin Hydrogels Prepared by Radical-Based Chain-Growth and Thiol-ene Mechanisms}, series = {Macromolecular Bioscience}, volume = {22}, journal = {Macromolecular Bioscience}, number = {4}, doi = {10.1002/mabi.202100274}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-318453}, year = {2022}, abstract = {Mucin, a high molecular mass hydrophilic glycoprotein, is the main component of mucus that coats every wet epithelium in animals. It is thus intrinsically biocompatible, and with its protein backbone and the o-glycosidic bound oligosaccharides, it contains a plethora of functional groups which can be used for further chemical modifications. Here, chain-growth and step-growth (thiol-ene) free-radical cross-linked hydrogels prepared from commercially available pig gastric mucin (PGM) are introduced and compared as cost-efficient and easily accessible alternative to the more broadly applied bovine submaxillary gland mucin. For this, PGM is functionalized with photoreactive acrylate groups or allyl ether moieties, respectively. Whereas homopolymerization of acrylate-functionalized polymers is performed, for thiol-ene cross-linking, the allyl-ether-functionalized PGM is cross-linked with thiol-functionalized hyaluronic acid. Morphology, mechanical properties, and cell compatibility of both kinds of PGM hydrogels are characterized and compared. Furthermore, the biocompatibility of these hydrogels can be evaluated in cell culture experiments.}, language = {en} } @article{FuchsKreczyBrueckneretal.2022, author = {Fuchs, Andreas and Kreczy, Dorothea and Br{\"u}ckner, Theresa and Gbureck, Uwe and Stahlhut, Philipp and Bengel, Melanie and Hoess, Andreas and Nies, Berthold and Bator, Julia and Klammert, Uwe and Linz, Christian and Ewald, Andrea}, title = {Bone regeneration capacity of newly developed spherical magnesium phosphate cement granules}, series = {Clinical Oral Investigations}, volume = {26}, journal = {Clinical Oral Investigations}, number = {3}, issn = {1436-3771}, doi = {10.1007/s00784-021-04231-w}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-268872}, pages = {2619-2633}, year = {2022}, abstract = {Objectives Magnesium phosphate-based cements begin to catch more attention as bone substitute materials and especially as alternatives for the more commonly used calcium phosphates. In bone substitutes for augmentation purposes, atraumatic materials with good biocompatibility and resorbability are favorable. In the current study, we describe the in vivo testing of novel bone augmentation materials in form of spherical granules based on a calcium-doped magnesium phosphate (CaMgP) cement. Materials and Methods Granules with diameters between 500 and 710 μm were fabricated via the emulsification of CaMgP cement pastes in a lipophilic liquid. As basic material, two different CaMgP formulations were used. The obtained granules were implanted into drill hole defects at the distal femoral condyle of 27 New Zealand white rabbits for 6 and 12 weeks. After explantation, the femora were examined via X-ray diffraction analysis, histological staining, radiological examination, and EDX measurement. Results Both granule types display excellent biocompatibility without any signs of inflammation and allow for proper bone healing without the interposition of connective tissue. CaMgP granules show a fast and continuous degradation and enable fully adequate bone regeneration. Conclusions Due to their biocompatibility, their degradation behavior, and their completely spherical morphology, these CaMgP granules present a promising bone substitute material for bone augmentation procedures, especially in sensitive areas. Clinical Relevance The mostly insufficient local bone supply after tooth extractions complicates prosthetic dental restoration or makes it even impossible. Therefore, bone augmentation procedures are oftentimes inevitable. Spherical CaMgP granules may represent a valuable bone replacement material in many situations.}, language = {en} } @article{BoehmStahlhutWeichholdetal.2022, author = {B{\"o}hm, Christoph and Stahlhut, Philipp and Weichhold, Jan and Hrynevich, Andrei and Teßmar, J{\"o}rg and Dalton, Paul D.}, title = {The Multiweek Thermal Stability of Medical-Grade Poly(ε-caprolactone) During Melt Electrowriting}, series = {Small}, volume = {18}, journal = {Small}, number = {3}, doi = {10.1002/smll.202104193}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-257741}, year = {2022}, abstract = {Melt electrowriting (MEW) is a high-resolution additive manufacturing technology that places unique constraints on the processing of thermally degradable polymers. With a single nozzle, MEW operates at low throughput and in this study, medical-grade poly(ε-caprolactone) (PCL) is heated for 25 d at three different temperatures (75, 85, and 95 °C), collecting daily samples. There is an initial increase in the fiber diameter and decrease in the jet speed over the first 5 d, then the MEW process remains stable for the 75 and 85 °C groups. When the collector speed is fixed to a value at least 10\% above the jet speed, the diameter remains constant for 25 d at 75 °C and only increases with time for 85 and 95 °C. Fiber fusion at increased layer height is observed for 85 and 95 °C, while the surface morphology of single fibers remain similar for all temperatures. The properties of the prints are assessed with no observable changes in the degree of crystallinity or the Young's modulus, while the yield strength decreases in later phases only for 95 °C. After the initial 5-d period, the MEW processing of PCL at 75 °C is extraordinarily stable with overall fiber diameters averaging 13.5 ± 1.0 µm over the entire 25-d period.}, language = {en} } @article{PereiraTrivanovićStahlhutetal.2022, author = {Pereira, Ana Rita and Trivanović, Drenka and Stahlhut, Philipp and Rudert, Maximilian and Groll, J{\"u}rgen and Herrmann, Marietta}, title = {Preservation of the na{\"i}ve features of mesenchymal stromal cells in vitro: Comparison of cell- and bone-derived decellularized extracellular matrix}, series = {Journal of Tissue Engineering}, volume = {13}, journal = {Journal of Tissue Engineering}, doi = {10.1177/20417314221074453}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-268835}, pages = {1-12}, year = {2022}, abstract = {The fate and behavior of bone marrow mesenchymal stem/stromal cells (BM-MSC) is bidirectionally influenced by their microenvironment, the stem cell niche, where a magnitude of biochemical and physical cues communicate in an extremely orchestrated way. It is known that simplified 2D in vitro systems for BM-MSC culture do not represent their na{\"i}ve physiological environment. Here, we developed four different 2D cell-based decellularized matrices (dECM) and a 3D decellularized human trabecular-bone scaffold (dBone) to evaluate BM-MSC behavior. The obtained cell-derived matrices provided a reliable tool for cell shape-based analyses of typical features associated with osteogenic differentiation at high-throughput level. On the other hand, exploratory proteomics analysis identified native bone-specific proteins selectively expressed in dBone but not in dECM models. Together with its architectural complexity, the physico-chemical properties of dBone triggered the upregulation of stemness associated genes and niche-related protein expression, proving in vitro conservation of the na{\"i}ve features of BM-MSC.}, language = {en} } @phdthesis{Stahlhut2023, author = {Stahlhut, Philipp}, title = {Konzeption und Aufbau einer Nanofokus Labor CT Anlage in Reflexionsgeometrie auf Basis eines Rasterelektronenmikroskops}, doi = {10.25972/OPUS-30264}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-302648}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2023}, abstract = {In der vorliegenden Arbeit werden die Konzeption und Realisierung eines Computertomographen zur Materialanalyse auf Basis eines Rasterelektronenmikroskops mit einem r{\"a}umlichen Aufl{\"o}sungsverm{\"o}gen im Nanometerbereich diskutiert. Durch einen fokussierten Elektronenstrahl, der mit einer Beschleunigungsspannung von 30 kV auf eine mikrostrukturierte Wolframnadel mit einem Spitzenradius von bis zu 50 nm gezielt wird, entsteht ein kleiner R{\"o}ntgenbrennfleck {\"u}ber den mit geometrischer Vergr{\"o}ßerung hochaufl{\"o}sende Projektionen eines zu untersuchenden Objekts erzeugt werden. Durch Rotation des Testobjekts werden Projektionen aus verschiedenen Blickwinkeln aufgenommen und {\"u}ber einen speziellen Rekonstruktionsalgorithmus zu einem 3-dimensionalen Bild zusammengef{\"u}gt. Bei der Beurteilung der Einzelkomponenten des Ger{\"a}ts wird insbesondere auf Struktur, Form und den elektrochemischen Herstellungsprozess der R{\"o}ntgenquelle eingegangen. Eine ausreichend genaue Positionierung von Messobjekt und R{\"o}ntgenbrennfleck wird {\"u}ber Piezoachsen realisiert, w{\"a}hrend die Stabilit{\"a}t des R{\"o}ntgenbrennflecks {\"u}ber die Elektronenoptik des Rasterelektronenmikroskops und die Form der Quellnadel optimiert wird. Das r{\"a}umliche Aufl{\"o}sungsverm{\"o}gen wird {\"u}ber die Linienspreizfunktion an Materialkanten abgesch{\"a}tzt. F{\"u}r eine Wolfram-Block-Quelle ergibt sich dabei ein Aufl{\"o}sungsverm{\"o}gen von 325 nm - 400 nm in 3D, w{\"a}hrend der Quellfleck einer Wolframnadel das Aufl{\"o}sungsverm{\"o}gen der Anlage auf 65 nm - 90 nm in 2D und 170 nm - 300 nm in 3D bei Messungen an einem AlCu29-Testobjekt anhebt. Außerdem werden die Auswirkungen der Phasenkontrastcharakteristik der R{\"o}ntgenquelle auf die rekonstruierten Bilder nach Anwendung eines Paganin-Filters diskutiert. Dabei zeigt sich, dass durch Anwendung des Filters ein verbessertes Signal-zu-Rausch-Verh{\"a}ltnis auf Kosten der r{\"a}umlichen Bildaufl{\"o}sung erzielt werden kann. Eine Vergleichsmessung mit einem kommerziell verf{\"u}gbaren R{\"o}ntgenmikroskop zeigt die St{\"a}rken des vorgestellten Systems bei Untersuchung von stark absorbierenden Messobjekten. Das kompakte Design erlaubt eine Weiterentwicklung in Richtung eines nanoCT-Moduls als Upgrade Option f{\"u}r Rasterelektronenmikroskope im Gegensatz zu den weitaus teureren bisher verbreiteten nanoCT-Ger{\"a}ten.}, subject = {Computertomographie}, language = {de} }