@article{PilusoVukicevieNoecheletal.2018, author = {Piluso, Susanna and Vukicevie, Radovan and N{\"o}chel, Ulrich and Braune, Steffen and Lendlein, Andreas and Neffe, Axel T.}, title = {Sequential alkyne-azide cycloadditions for functionalized gelatin hydrogel formation}, series = {European polymer journal}, volume = {100}, journal = {European polymer journal}, publisher = {Elsevier}, address = {Oxford}, issn = {0014-3057}, doi = {10.1016/j.eurpolymj.2018.01.017}, pages = {77 -- 85}, year = {2018}, abstract = {While click chemistry reactions for biopolymer network formation are attractive as the defined reactions may allow good control of the network formation and enable subsequent functionalization, tailoring of gelatin network properties over a wide range of mechanical properties has yet to be shown. Here, it is demonstrated that copper-catalyzed alkyne-azide cycloaddition of alkyne functionalized gelatin with diazides gave hydrogel networks with properties tailorable by the ratio of diazide to gelatin and diazide rigidity. 4,4′-diazido-2,2′-stilbenedisulfonic acid, which has been used as rigid crosslinker, yielded hydrogels with Young's moduli E of 50-390 kPa and swelling degrees Q of 150-250 vol.\%, while the more flexible 1,8-diazidooctane resulted in hydrogels with E = 125-280 kPa and Q = 225-470 vol.\%. Storage moduli could be varied by two orders of magnitude (G′ = 100-20,000 Pa). An indirect cytotoxicity test did not show cytotoxic properties. Even when employing 1:1 ratios of alkyne and azide moieties, the hydrogels were shown to contain both, unreacted alkyne groups on the gelatin backbone as well as dangling chains carrying azide groups as shown by reaction with functionalized fluorescein. The free groups, which can be tailored by the employed ratio of the reactants, are accessible for covalent attachment of drugs, as was demonstrated by functionalization with dexamethasone. The sequential network formation and functionalization with click chemistry allows access to multifunctional materials relevant for medical applications.}, language = {en} } @phdthesis{Tunn2020, author = {Tunn, Isabell}, title = {From single molecules to bulk materials: tuning the viscoelastic properties of coiled coil cross-linked hydrogels}, doi = {10.25932/publishup-47595}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-475955}, school = {Universit{\"a}t Potsdam}, pages = {XVI, 140}, year = {2020}, abstract = {The development of bioinspired self-assembling materials, such as hydrogels, with promising applications in cell culture, tissue engineering and drug delivery is a current focus in material science. Biogenic or bioinspired proteins and peptides are frequently used as versatile building blocks for extracellular matrix (ECM) mimicking hydrogels. However, precisely controlling and reversibly tuning the properties of these building blocks and the resulting hydrogels remains challenging. Precise control over the viscoelastic properties and self-healing abilities of hydrogels are key factors for developing intelligent materials to investigate cell matrix interactions. Thus, there is a need to develop building blocks that are self-healing, tunable and self-reporting. This thesis aims at the development of α-helical peptide building blocks, called coiled coils (CCs), which integrate these desired properties. Self-healing is a direct result of the fast self-assembly of these building blocks when used as material cross-links. Tunability is realized by means of reversible histidine (His)-metal coordination bonds. Lastly, implementing a fluorescent readout, which indicates the CC assembly state, self-reporting hydrogels are obtained. Coiled coils are abundant protein folding motifs in Nature, which often have mechanical function, such as in myosin or fibrin. Coiled coils are superhelices made up of two or more α-helices wound around each other. The assembly of CCs is based on their repetitive sequence of seven amino acids, so-called heptads (abcdefg). Hydrophobic amino acids in the a and d position of each heptad form the core of the CC, while charged amino acids in the e and g position form ionic interactions. The solvent-exposed positions b, c and f are excellent targets for modifications since they are more variable. His-metal coordination bonds are strong, yet reversible interactions formed between the amino acid histidine and transition metal ions (e.g. Ni2+, Cu2+ or Zn2+). His-metal coordination bonds essentially contribute to the mechanical stability of various high-performance proteinaceous materials, such as spider fangs, Nereis worm jaws and mussel byssal threads. Therefore, I bioengineered reversible His-metal coordination sites into a well-characterized heterodimeric CC that served as tunable material cross-link. Specifically, I took two distinct approaches facilitating either intramolecular (Chapter 4.2) and/or intermolecular (Chapter 4.3) His-metal coordination. Previous research suggested that force-induced CC unfolding in shear geometry starts from the points of force application. In order to tune the stability of a heterodimeric CC in shear geometry, I inserted His in the b and f position at the termini of force application (Chapter 4.2). The spacing of His is such that intra-CC His-metal coordination bonds can form to bridge one helical turn within the same helix, but also inter-CC coordination bonds are not generally excluded. Starting with Ni2+ ions, Raman spectroscopy showed that the CC maintained its helical structure and the His residues were able to coordinate Ni2+. Circular dichroism (CD) spectroscopy revealed that the melting temperature of the CC increased by 4 °C in the presence of Ni2+. Using atomic force microscope (AFM)-based single molecule force spectroscopy, the energy landscape parameters of the CC were characterized in the absence and the presence of Ni2+. His-Ni2+ coordination increased the rupture force by ~10 pN, accompanied by a decrease of the dissociation rate constant. To test if this stabilizing effect can be transferred from the single molecule level to the bulk viscoelastic material properties, the CC building block was used as a non-covalent cross-link for star-shaped poly(ethylene glycol) (star-PEG) hydrogels. Shear rheology revealed a 3-fold higher relaxation time in His-Ni2+ coordinating hydrogels compared to the hydrogel without metal ions. This stabilizing effect was fully reversible when using an excess of the metal chelator ethylenediaminetetraacetate (EDTA). The hydrogel properties were further investigated using different metal ions, i.e. Cu2+, Co2+ and Zn2+. Overall, these results suggest that Ni2+, Cu2+ and Co2+ primarily form intra-CC coordination bonds while Zn2+ also participates in inter-CC coordination bonds. This may be a direct result of its different coordination geometry. Intermolecular His-metal coordination bonds in the terminal regions of the protein building blocks of mussel byssal threads are primarily formed by Zn2+ and were found to be intimately linked to higher-order assembly and self-healing of the thread. In the above example, the contribution of intra-CC and inter-CC His-Zn2+ cannot be disentangled. In Chapter 4.3, I redesigned the CC to prohibit the formation of intra-CC His-Zn2+ coordination bonds, focusing only on inter-CC interactions. Specifically, I inserted His in the solvent-exposed f positions of the CC to focus on the effect of metal-induced higher-order assembly of CC cross-links. Raman and CD spectroscopy revealed that this CC building block forms α-helical Zn2+ cross-linked aggregates. Using this CC as a cross-link for star-PEG hydrogels, I showed that the material properties can be switched from viscoelastic in the absence of Zn2+ to elastic-like in the presence of Zn2+. Moreover, the relaxation time of the hydrogel was tunable over three orders of magnitude when using different Zn2+:His ratios. This tunability is attributed to a progressive transformation of single CC cross-links into His-Zn2+ cross-linked aggregates, with inter-CC His-Zn2+ coordination bonds serving as an additional, cross-linking mode. Rheological characterization of the hydrogels with inter-CC His-Zn2+ coordination raised the question whether the His-Zn2+ coordination bonds between CCs or also the CCs themselves rupture when shear strain is applied. In general, the amount of CC cross-links initially formed in the hydrogel as well as the amount of CC cross-links breaking under force remains to be elucidated. In order to more deeply probe these questions and monitor the state of the CC cross-links when force is applied, a fluorescent reporter system based on F{\"o}rster resonance energy transfer (FRET) was introduced into the CC (Chapter 4.4). For this purpose, the donor-acceptor pair carboxyfluorescein and tetramethylrhodamine was used. The resulting self-reporting CC showed a FRET efficiency of 77 \% in solution. Using this fluorescently labeled CC as a self-reporting, reversible cross-link in an otherwise covalently cross-linked star-PEG hydrogel enabled the detection of the FRET efficiency change under compression force. This proof-of-principle result sets the stage for implementing the fluorescently labeled CCs as molecular force sensors in non-covalently cross-linked hydrogels. In summary, this thesis highlights that rationally designed CCs are excellent reversibly tunable, self-healing and self-reporting hydrogel cross-links with high application potential in bioengineering and biomedicine. For the first time, I demonstrated that His-metal coordination-based stabilization can be transferred from the single CC level to the bulk material with clear viscoelastic consequences. Insertion of His in specific sequence positions was used to implement a second non-covalent cross-linking mode via intermolecular His-metal coordination. This His-metal binding induced aggregation of the CCs enabled for reversibly tuning the hydrogel properties from viscoelastic to elastic-like. As a proof-of-principle to establish self-reporting CCs as material cross-links, I labeled a CC with a FRET pair. The fluorescently labelled CC acts as a molecular force sensor and first preliminary results suggest that the CC enables the detection of hydrogel cross-link failure under compression force. In the future, fluorescently labeled CC force sensors will likely not only be used as intelligent cross-links to study the failure of hydrogels but also to investigate cell-matrix interactions in 3D down to the single molecule level.}, language = {en} } @article{PilusoLendleinNeffe2017, author = {Piluso, Susanna and Lendlein, Andreas and Neffe, Axel T.}, title = {Enzymatic action as switch of bulk to surface degradation of clicked gelatin-based networks}, series = {Polymers for advanced technologies}, volume = {28}, journal = {Polymers for advanced technologies}, publisher = {Wiley}, address = {Hoboken}, issn = {1042-7147}, doi = {10.1002/pat.3962}, pages = {1318 -- 1324}, year = {2017}, abstract = {Polymer degradation occurs under physiological conditions in vitro and in vivo, especially when bonds susceptible to hydrolysis are present in the polymer. Understanding of the degradation mechanism, changes of material properties over time, and overall rate of degradation is a necessary prerequisite for the knowledge-based design of polymers with applications in biomedicine. Here, hydrolytic degradation studies of gelatin-based networks synthesized by copper-catalyzed azide-alkyne cycloaddition reaction are reported, which were performed with or without addition of an enzyme. In all cases, networks with a stilbene as crosslinker proofed to be more resistant to degradation than when an octyl diazide was used. Without addition of an enzyme, the rate of degradation was ruled by the crosslinking density of the network and proceeded via a bulk degradation mechanism. Addition of Clostridium histolyticum collagenase resulted in a much enhanced rate of degradation, which furthermore occurred via surface erosion. The mesh size of the hydrogels (>7nm) was in all cases larger than the hydrodynamic radius of the enzyme (4.5nm) so that even in very hydrophilic networks with large mesh size enzymes may be used to induce a fast surface degradation mechanism. This observation is of general interest when designing hydrogels to be applied in the presence of enzymes, as the degradation mechanism and material performance are closely interlinked. Copyright (c) 2016 John Wiley \& Sons, Ltd.}, language = {en} } @phdthesis{Vacogne2016, author = {Vacogne, Charlotte D.}, title = {New synthetic routes towards well-defined polypeptides, morphologies and hydrogels}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-396366}, school = {Universit{\"a}t Potsdam}, pages = {xii, 175}, year = {2016}, abstract = {Proteins are natural polypeptides produced by cells; they can be found in both animals and plants, and possess a variety of functions. One of these functions is to provide structural support to the surrounding cells and tissues. For example, collagen (which is found in skin, cartilage, tendons and bones) and keratin (which is found in hair and nails) are structural proteins. When a tissue is damaged, however, the supporting matrix formed by structural proteins cannot always spontaneously regenerate. Tailor-made synthetic polypeptides can be used to help heal and restore tissue formation. Synthetic polypeptides are typically synthesized by the so-called ring opening polymerization (ROP) of α-amino acid N-carboxyanhydrides (NCA). Such synthetic polypeptides are generally non-sequence-controlled and thus less complex than proteins. As such, synthetic polypeptides are rarely as efficient as proteins in their ability to self-assemble and form hierarchical or structural supramolecular assemblies in water, and thus, often require rational designing. In this doctoral work, two types of amino acids, γ-benzyl-L/D-glutamate (BLG / BDG) and allylglycine (AG), were selected to synthesize a series of (co)polypeptides of different compositions and molar masses. A new and versatile synthetic route to prepare polypeptides was developed, and its mechanism and kinetics were investigated. The polypeptide properties were thoroughly studied and new materials were developed from them. In particular, these polypeptides were able to aggregate (or self-assemble) in solution into microscopic fibres, very similar to those formed by collagen. By doing so, they formed robust physical networks and organogels which could be processed into high water-content, pH-responsive hydrogels. Particles with highly regular and chiral spiral morphologies were also obtained by emulsifying these polypeptides. Such polypeptides and the materials derived from them are, therefore, promising candidates for biomedical applications.}, language = {en} } @article{AdelsbergerBivigouKoumbaMiasnikovaetal.2015, author = {Adelsberger, Joseph and Bivigou Koumba, Achille Mayelle and Miasnikova, Anna and Busch, Peter and Laschewsky, Andr{\´e} and M{\"u}ller-Buschbaum, Peter and Papadakis, Christine M.}, title = {Polystyrene-block-poly (methoxy diethylene glycol acrylate)-block-polystyrene triblock copolymers in aqueous solution-a SANS study of the temperature-induced switching behavior}, series = {Colloid and polymer science : official journal of the Kolloid-Gesellschaft}, volume = {293}, journal = {Colloid and polymer science : official journal of the Kolloid-Gesellschaft}, number = {5}, publisher = {Springer}, address = {New York}, issn = {0303-402X}, doi = {10.1007/s00396-015-3535-6}, pages = {1515 -- 1523}, year = {2015}, abstract = {A concentrated solution of a symmetric triblock copolymer with a thermoresponsive poly(methoxy diethylene glycol acrylate) (PMDEGA) middle block and short hydrophobic, fully deuterated polystyrene end blocks is investigated in D2O where it undergoes a lower critical solution temperature-type phase transition at ca. 36 A degrees C. Small-angle neutron scattering (SANS) in a wide temperature range (15-50 A degrees C) is used to characterize the size and inner structure of the micelles as well as the correlation between the micelles and the formation of aggregates by the micelles above the cloud point (CP). A model featuring spherical core-shell micelles, which are correlated by a hard-sphere potential or a sticky hard-sphere potential together with a Guinier form factor describing aggregates formed by the micelles above the CP, fits the SANS curves well in the entire temperature range. The thickness of the thermoresponsive micellar PMDEGA shell as well as the hard-sphere radius increase slightly already below the cloud point. Whereas the thickness of the thermoresponsive micellar shell hardly shrinks when heating through the CP and up to 50 A degrees C, the hard-sphere radius decreases within 3.5 K at the CP. The volume fraction decreases already significantly below the CP, which may be at the origin of the previously observed gel-sol transition far below the CP (Miasnikova et al., Langmuir 28: 4479-4490, 2012). Above the CP, small, and at higher temperatures, large aggregates are formed by the micelles.}, language = {en} } @article{RichterZakrevskyyEiseleetal.2014, author = {Richter, Marcel and Zakrevskyy, Yuriy and Eisele, Michael and Lomadze, Nino and Santer, Svetlana and von Klitzing, Regine}, title = {Effect of pH, co-monomer content, and surfactant structure on the swelling behavior of microgel-azobenzene-containing surfactant complex}, series = {Polymer : the international journal for the science and technology of polymers}, volume = {55}, journal = {Polymer : the international journal for the science and technology of polymers}, number = {25}, publisher = {Elsevier}, address = {Oxford}, issn = {0032-3861}, doi = {10.1016/j.polymer.2014.10.027}, pages = {6513 -- 6518}, year = {2014}, abstract = {The contraction/swelling transition of anionic PNIPAM-co-AAA particles can be manipulated by light using interactions with cationic azobenzene-containing surfactant. In this study the influence of pH-buffers and their concentrations, the charge density (AAA content) in microgel particles as well as the spacer length of the surfactant on the complex formation between the microgel and surfactant is investigated. It is shown that the presence of pH buffer can lead to complete blocking of the interactions in such complexes and the resulting microgel contraction/swelling response. There is a clear competition between the buffer ions and the surfactant molecules interacting with microgel particles. When working in pure water solutions with fixed concentration (charge density) of microgel, the contraction/swelling of the particles is controlled only by relative concentration (charge ratio) of the surfactant and AAA groups of the microgel. Furthermore, the particle contraction is more efficient for shorter spacer length of the surfactant. The onset point of the contraction process is not affected by the surfactant hydrophobicity. This work provides new insight into the interaction between microgel particles and photo-sensitive surfactants, which offers high potential in new sensor systems. (C) 2014 Elsevier Ltd. All rights reserved.}, language = {en} } @article{PilusoHieblGorbetal.2011, author = {Piluso, Susanna and Hiebl, Bernhard and Gorb, Stanislav N. and Kovalev, Alexander and Lendlein, Andreas and Neffe, Axel T.}, title = {Hyaluronic acid-based hydrogels crosslinked by copper-catalyzed azide-alkyne cycloaddition with tailorable mechanical properties}, series = {The international journal of artificial organs}, volume = {34}, journal = {The international journal of artificial organs}, number = {2}, publisher = {Wichtig}, address = {Milano}, issn = {0391-3988}, doi = {10.5301/IJAO.2011.6394}, pages = {192 -- 197}, year = {2011}, abstract = {Biopolymers of the extracellular matrix are attractive starting materials for providing degradable and biocompatible biomaterials. In this study, hyaluronic acid-based hydrogels with tunable mechanical properties were prepared by the use of copper-catalyzed azide-alkyne cycloaddition (known as "click chemistry"). Alkyne-functionalized hyaluronic acid was crosslinked with linkers having two terminal azide functionalities, varying crosslinker density as well as the lengths and rigidity of the linker molecules. By variation of the crosslinker density and crosslinker type, hydrogels with elastic moduli in the range of 0.5-4 kPa were prepared. The washed materials contained a maximum of 6.8 mg copper per kg dry weight and the eluate of the gel crosslinked with diazidostilbene did not show toxic effects on L929 cells. The hyaluronic acid-based hydrogels have potential as biomaterials for cell culture or soft tissue regeneration applications.}, language = {en} } @article{NeffeLoebusZaupaetal.2011, author = {Neffe, Axel T. and Loebus, Axel and Zaupa, Alessandro and St{\"o}tzel, Christian and M{\"u}ller, Frank A. and Lendlein, Andreas}, title = {Gelatin functionalization with tyrosine derived moieties to increase the interaction with hydroxyapatite fillers}, series = {Acta biomaterialia}, volume = {7}, journal = {Acta biomaterialia}, number = {4}, publisher = {Elsevier}, address = {Oxford}, issn = {1742-7061}, doi = {10.1016/j.actbio.2010.11.025}, pages = {1693 -- 1701}, year = {2011}, abstract = {Combining gelatins functionalized with the tyrosine-derived groups desaminotyrosine or desaminotyrosyl tyrosine with hydroxyapatite (HAp) led to the formation of composite materials with much lower swelling ratios than those of the pure matrices. Shifts of the infra-red (IR) bands related to the free carboxyl groups could be observed in the presence of HAp, which suggested a direct interaction of matrix and filler that formed additional physical cross-links in the material. In tensile tests and rheological measurements the composites equilibrated in water had increased Young's moduli (from 200 kPa up to 2 MPa) and tensile strengths (from 57 kPa up to 1.1 MPa) compared with the matrix polymers without affecting the elongation at break. Furthermore, an increased thermal stability of the networks from 40 to 85 degrees C could be demonstrated. The differences in the behaviour of the functionalized gelatins compared with pure gelatin as a matrix suggested an additional stabilizing bond between the incorporated aromatic groups and the HAp as supported by the IR results. The composites can potentially be applied as bone fillers.}, language = {en} } @article{ZhongWangAdelsbergeretal.2011, author = {Zhong, Qi and Wang, Weinan and Adelsberger, Joseph and Golosova, Anastasia and Koumba, Achille M. Bivigou and Laschewsky, Andr{\´e} and Funari, Sergio S. and Perlich, Jan and Roth, Stephan V. and Papadakis, Christine M. and M{\"u}ller-Buschbaum, Peter}, title = {Collapse transition in thin films of poly(methoxydiethylenglycol acrylate)}, series = {Colloid and polymer science : official journal of the Kolloid-Gesellschaft}, volume = {289}, journal = {Colloid and polymer science : official journal of the Kolloid-Gesellschaft}, number = {5-6}, publisher = {Springer}, address = {New York}, issn = {0303-402X}, doi = {10.1007/s00396-011-2384-1}, pages = {569 -- 581}, year = {2011}, abstract = {The thermal behavior of poly(methoxydiethylenglycol acrylate) (PMDEGA) is studied in thin hydrogel films on solid supports and is compared with the behavior in aqueous solution. The PMDEGA hydrogel film thickness is varied from 2 to 422 nm. Initially, these films are homogenous, as measured with optical microscopy, atomic force microscopy, X-ray reflectivity, and grazing-incidence small-angle X-ray scattering (GISAXS). However, they tend to de-wet when stored under ambient conditions. Along the surface normal, no long-ranged correlations between substrate and film surface are detected with GISAXS, due to the high mobility of the polymer at room temperature. The swelling of the hydrogel films as a function of the water vapor pressure and the temperature are probed for saturated water vapor pressures between 2,380 and 3,170 Pa. While the swelling capability is found to increase with water vapor pressure, swelling in dependence on the temperature revealed a collapse phase transition of a lower critical solution temperature type. The transition temperature decreases from 40.6 A degrees C to 36.6 A degrees C with increasing film thickness, but is independent of the thickness for very thin films below a thickness of 40 nm. The observed transition temperature range compares well with the cloud points observed in dilute (0.1 wt.\%) and semi-dilute (5 wt.\%) solution which decrease from 45 A degrees C to 39 A degrees C with increasing concentration.}, language = {en} } @article{TartivelBehlSchroeteretal.2012, author = {Tartivel, Lucile and Behl, Marc and Schr{\"o}ter, Michael and Lendlein, Andreas}, title = {Hydrogel networks based on ABA triblock copolymers}, series = {Journal of applied biomaterials \& functional materials}, volume = {10}, journal = {Journal of applied biomaterials \& functional materials}, number = {3}, publisher = {Wichtig}, address = {Milano}, issn = {2280-8000}, doi = {10.5301/JABFM.2012.10295}, pages = {243 -- 248}, year = {2012}, abstract = {Background: Triblock copolymers from hydrophilic oligo(ethylene glycol) segment A and oligo(propylene glycol) segment B, providing an ABA structure (OEG-OPG-OEG triblock), are known to be biocompatible and are used as self-solidifying gels in drug depots. A complete removal of these depots would be helpful in cases of undesired side effects of a drug, but this remains a challenge as they liquefy below their transition temperature. Therefore we describe the synthesis of covalently cross-linked hydrogel networks. Method: Triblock copolymer-based hydrogels were created by irradiating aqueous solutions of the corresponding macro-dimethacrylates with UV light. The degree of swelling, swelling kinetics, mechanical properties and morphology of the networks were investigated. Results: Depending on precursor concentration, equilibrium degree of swelling of the films ranged between 500\% and 880\% and was reached in 1 hour. In addition, values for storage and loss moduli of the hydrogel networks were in the 100 Pa to 10 kPa range. Conclusion: Although OEG-OPG-OEG triblocks are known for their micellization, which could hamper polymer network formation, reactive OEG-OPG-OEG triblock oligomers could be successfully polymerized into hydrogel networks. The degree of swelling of these hydrogels depends on their molecular weight and on the oligomer concentration used for hydrogel preparation. In combination with the temperature sensitivity of the ABA triblock copolymers, it is assumed that such hydrogels might be beneficial for future medical applications -e.g., removable drug release systems.}, language = {en} }