TY - JOUR A1 - Wieland, Annalena A1 - Strissel, Pamela L. A1 - Schorle, Hannah A1 - Bakirci, Ezgi A1 - Janzen, Dieter A1 - Beckmann, Matthias W. A1 - Eckstein, Markus A1 - Dalton, Paul D. A1 - Strick, Reiner T1 - Brain and breast cancer cells with PTEN loss of function reveal enhanced durotaxis and RHOB dependent amoeboid migration utilizing 3D scaffolds and aligned microfiber tracts JF - Cancers N2 - Background: Glioblastoma multiforme (GBM) and metastatic triple-negative breast cancer (TNBC) with PTEN mutations often lead to brain dissemination with poor patient outcome, thus new therapeutic targets are needed. To understand signaling, controlling the dynamics and mechanics of brain tumor cell migration, we implemented GBM and TNBC cell lines and designed 3D aligned microfibers and scaffolds mimicking brain structures. Methods: 3D microfibers and scaffolds were printed using melt electrowriting. GBM and TNBC cell lines with opposing PTEN genotypes were analyzed with RHO-ROCK-PTEN inhibitors and PTEN rescue using live-cell imaging. RNA-sequencing and qPCR of tumor cells in 3D with microfibers were performed, while scanning electron microscopy and confocal microscopy addressed cell morphology. Results: In contrast to the PTEN wildtype, GBM and TNBC cells with PTEN loss of function yielded enhanced durotaxis, topotaxis, adhesion, amoeboid migration on 3D microfibers and significant high RHOB expression. Functional studies concerning RHOB-ROCK-PTEN signaling confirmed the essential role for the above cellular processes. Conclusions: This study demonstrates a significant role of the PTEN genotype and RHOB expression for durotaxis, adhesion and migration dependent on 3D. GBM and TNBC cells with PTEN loss of function have an affinity for stiff brain structures promoting metastasis. 3D microfibers represent an important tool to model brain metastasizing tumor cells, where RHO-inhibitors could play an essential role for improved therapy. KW - 3D tumor model KW - 3D microfiber KW - amoeboid cell migration KW - brain cancer KW - breast cancer KW - PTEN KW - RHO KW - ROCK KW - durotaxis KW - topotaxis Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-248443 SN - 2072-6694 VL - 13 IS - 20 ER - TY - JOUR A1 - Weigand, Annika A1 - Boos, Anja M. A1 - Tasbihi, Kereshmeh A1 - Beier, Justus P. A1 - Dalton, Paul D. A1 - Schrauder, Michael A1 - Horch, Raymund E. A1 - Beckmann, Matthias W. A1 - Strissel, Pamela L. A1 - Strick, Reiner T1 - Selective isolation and characterization of primary cells from normal breast and tumors reveal plasticity of adipose derived stem cells JF - Breast Cancer Research N2 - Background There is a need to establish more cell lines from breast tumors in contrast to immortalized cell lines from metastatic effusions in order to represent the primary tumor and not principally metastatic biology of breast cancer. This investigation describes the simultaneous isolation, characterization, growth and function of primary mammary epithelial cells (MEC), mesenchymal cells (MES) and adipose derived stem cells (ADSC) from four normal breasts, one inflammatory and one triple-negative ductal breast tumors. Methods A total of 17 cell lines were established and gene expression was analyzed for MEC and MES (n = 42) and ADSC (n = 48) and MUC1, pan-KRT, CD90 and GATA-3 by immunofluorescence. DNA fingerprinting to track cell line identity was performed between original primary tissues and isolates. Functional studies included ADSC differentiation, tumor MES and MEC invasion co-cultured with ADSC-conditioned media (CM) and MES adhesion and growth on 3D-printed scaffolds. Results Comparative analysis showed higher gene expression of EPCAM, CD49f, CDH1 and KRTs for normal MEC lines; MES lines e.g. Vimentin, CD10, ACTA2 and MMP9; and ADSC lines e.g. CD105, CD90, CDH2 and CDH11. Compared to the mean of all four normal breast cell lines, both breast tumor cell lines demonstrated significantly lower ADSC marker gene expression, but higher expression of mesenchymal and invasion gene markers like SNAI1 and MMP2. When compared with four normal ADSC differentiated lineages, both tumor ADSC showed impaired osteogenic and chondrogenic but enhanced adipogenic differentiation and endothelial-like structures, possibly due to high PDGFRB and CD34. Addressing a functional role for overproduction of adipocytes, we initiated 3D-invasion studies including different cell types from the same patient. CM from ADSC differentiating into adipocytes induced tumor MEC 3D-invasion via EMT and amoeboid phenotypes. Normal MES breast cells adhered and proliferated on 3D-printed scaffolds containing 20 fibers, but not on 2.5D-printed scaffolds with single fiber layers, important for tissue engineering. Conclusion Expression analyses confirmed successful simultaneous cell isolations of three different phenotypes from normal and tumor primary breast tissues. Our cell culture studies support that breast-tumor environment differentially regulates tumor ADSC plasticity as well as cell invasion and demonstrates applications for regenerative medicine. KW - Normal breast KW - Breast cancer KW - Stem cells plasticity KW - Primary cell lines KW - Tissue engineering Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-164759 VL - 18 IS - 32 ER - TY - JOUR A1 - Wang, Shuang A1 - Sarwat, Mariah A1 - Wang, Peng A1 - Surrao, Denver C. A1 - Harkin, Damien G. A1 - St John, James A. A1 - Bolle, Eleonore C. L. A1 - Forget, Aurelien A1 - Dalton, Paul D. A1 - Dargaville, Tim R. T1 - Hydrogels with Cell Adhesion Peptide‐Decorated Channel Walls for Cell Guidance JF - Macromolecular Rapid Communications N2 - A method is reported for making hollow channels within hydrogels decorated with cell‐adhesion peptides exclusively at the channel surface. Sacrificial fibers of different diameters are used to introduce channels within poly(ethylene glycol) hydrogels crosslinked with maleimide‐thiol chemistry, which are backfilled with a cysteine‐containing peptide solution which is conjugated to the lumen with good spatial efficiency. This allows for peptide patterning in only the areas of the hydrogel where they are needed when used as cell‐guides, reducing the amount of required peptide 20‐fold when compared to bulk functionalization. The power of this approach is highlighted by successfully using these patterned hydrogels without active perfusion to guide fibroblasts and olfactory ensheathing cells—the latter having unique potential in neural repair therapies. KW - 3D printing KW - cell guidance KW - cell transplantation KW - melt electrowriting KW - synthetic hydrogels Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-218031 VL - 41 IS - 15 ER - TY - JOUR A1 - Thibaudeau, Laure A1 - Taubenberger, Anna V. A1 - Holzapfel, Boris M. A1 - Quent, Verena M. A1 - Fuehrmann, Tobias A1 - Hesami, Parisa A1 - Brown, Toby D. A1 - Dalton, Paul D. A1 - Power, Carl A. A1 - Hollier, Brett G. A1 - Hutmacher, Dietmar W. T1 - A tissue-engineered humanized xenograft model of human breast cancer metastasis to bone JF - Disease Models & Mechanisms N2 - The skeleton is a preferred homing site for breast cancer metastasis. To date, treatment options for patients with bone metastases are mostly palliative and the disease is still incurable. Indeed, key mechanisms involved in breast cancer osteotropism are still only partially understood due to the lack of suitable animal models to mimic metastasis of human tumor cells to a human bone microenvironment. In the presented study, we investigate the use of a human tissue-engineered bone construct to develop a humanized xenograft model of breast cancer-induced bone metastasis in a murine host. Primary human osteoblastic cell-seeded melt electrospun scaffolds in combination with recombinant human bone morphogenetic protein 7 were implanted subcutaneously in non-obese diabetic/severe combined immunodeficient mice. The tissue-engineered constructs led to the formation of a morphologically intact 'organ' bone incorporating a high amount of mineralized tissue, live osteocytes and bone marrow spaces. The newly formed bone was largely humanized, as indicated by the incorporation of human bone cells and human-derived matrix proteins. After intracardiac injection, the dissemination of luciferase-expressing human breast cancer cell lines to the humanized bone ossicles was detected by bioluminescent imaging. Histological analysis revealed the presence of metastases with clear osteolysis in the newly formed bone. Thus, human tissue-engineered bone constructs can be applied efficiently as a target tissue for human breast cancer cells injected into the blood circulation and replicate the osteolytic phenotype associated with breast cancer-induced bone lesions. In conclusion, we have developed an appropriate model for investigation of species-specific mechanisms of human breast cancer-related bone metastasis in vivo. KW - breast cancer KW - bone metastasis KW - humanized xenograft model KW - melt electrospinning KW - tissue engineering KW - osteotropism KW - in vivo KW - stem-cell niche KW - human prostate-cancer KW - morphogenetic protein KW - osteoprogenitor cells KW - endochondral ossification KW - mouse model KW - trabecular bone KW - calcium phosphate KW - skeletal metastases Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-117466 VL - 7 IS - 2 ER - TY - INPR A1 - Schaefer, Natascha A1 - Janzen, Dieter A1 - Bakirci, Ezgi A1 - Hrynevich, Andrei A1 - Dalton, Paul D. A1 - Villmann, Carmen T1 - 3D Electrophysiological Measurements on Cells Embedded within Fiber-Reinforced Matrigel T2 - Advanced Healthcare Materials N2 - 2D electrophysiology is often used to determine the electrical properties of neurons, while in the brain, neurons form extensive 3D networks. Thus, performing electrophysiology in a 3D environment provides a closer situation to the physiological condition and serves as a useful tool for various applications in the field of neuroscience. In this study, we established 3D electrophysiology within a fiber-reinforced matrix to enable fast readouts from transfected cells, which are often used as model systems for 2D electrophysiology. Using melt electrowriting (MEW) of scaffolds to reinforce Matrigel, we performed 3D electrophysiology on a glycine receptor-transfected Ltk-11 mouse fibroblast cell line. The glycine receptor is an inhibitory ion channel associated when mutated with impaired neuromotor behaviour. The average thickness of the MEW scaffold was 141.4 ± 5.7µm, using 9.7 ± 0.2µm diameter fibers, and square pore spacings of 100 µm, 200 µm and 400 µm. We demonstrate, for the first time, the electrophysiological characterization of glycine receptor-transfected cells with respect to agonist efficacy and potency in a 3D matrix. With the MEW scaffold reinforcement not interfering with the electrophysiology measurement, this approach can now be further adapted and developed for different kinds of neuronal cultures to study and understand pathological mechanisms under disease conditions. KW - 3D cultures Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-244194 ER - TY - JOUR A1 - Robinson, Thomas M. A1 - Hutmacher, Dietmar W. A1 - Dalton, Paul D. T1 - The next frontier in melt electrospinning: taming the jet JF - Advanced Functional Materials N2 - There is a specialized niche for the electrohydrodynamic jetting of melts, from biomedical products to filtration and soft matter applications. The next frontier includes optics, microfluidics, flexible electronic devices, and soft network composites in biomaterial science and soft robotics. The recent emphasis on reproducibly direct‐writing continual molten jets has enabled a spectrum of contemporary microscale 3D objects to be fabricated. One strong suit of melt processing is the capacity for the jet to solidify rapidly into a fiber, thus fixing a particular structure into position. The ability to direct‐write complex and multiscaled architectures and structures has greatly contributed to a large number of recent studies, explicitly, toward fiber–hydrogel composites and fugitive inks, and has expanded into several biomedical applications such as cartilage, skin, periosteum, and cardiovascular tissue engineering. Following the footsteps of a publication that summarized melt electrowriting literature up to 2015, the most recent literature from then until now is reviewed to provide a continuous and comprehensive timeline that demonstrates the latest advances as well as new perspectives for this emerging technology. KW - 3D printing KW - additive manufacturing KW - eletrhydrodynamic KW - melt electrospinning writing Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-204819 VL - 29 ER - TY - JOUR A1 - Mieszczanek, Pawel A1 - Robinson, Thomas M. A1 - Dalton, Paul D. A1 - Hutmacher, Dietmar W. T1 - Convergence of Machine Vision and Melt Electrowriting JF - Advanced Materials N2 - Melt electrowriting (MEW) is a high-resolution additive manufacturing technology that balances multiple parametric variables to arrive at a stable fabrication process. The better understanding of this balance is underscored here using high-resolution camera vision of jet stability profiles in different electrical fields. Complementing this visual information are fiber-diameter measurements obtained at precise points, allowing the correlation to electrified jet properties. Two process signatures—the jet angle and for the first time, the Taylor cone area—are monitored and analyzed with a machine vision system, while SEM imaging for diameter measurement correlates real-time information. This information, in turn, allows the detection and correction of fiber pulsing for accurate jet placement on the collector, and the in-process assessment of the fiber diameter. Improved process control is used to successfully fabricate collapsible MEW tubes; structures that require exceptional accuracy and printing stability. Using a precise winding angle of 60° and 300 layers, the resulting 12 mm-thick tubular structures have elastic snap-through instabilities associated with mechanical metamaterials. This study provides a detailed analysis of the fiber pulsing occurrence in MEW and highlights the importance of real-time monitoring of the Taylor cone volume to better understand, control, and predict printing instabilities. KW - polycaprolactone KW - 3D printing KW - digitization KW - electrohydrodynamic KW - melt electrospinning writing Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-256365 VL - 33 IS - 29 ER - TY - JOUR A1 - Mechau, Jannik A1 - Frank, Andreas A1 - Bakirci, Ezgi A1 - Gumbel, Simon A1 - Jungst, Tomasz A1 - Giesa, Reiner A1 - Groll, Jürgen A1 - Dalton, Paul D. A1 - Schmidt, Hans‐Werner T1 - Hydrophilic (AB)\(_{n}\) Segmented Copolymers for Melt Extrusion‐Based Additive Manufacturing JF - Macromolecular Chemistry and Physics N2 - Several manufacturing technologies beneficially involve processing from the melt, including extrusion‐based printing, electrospinning, and electrohydrodynamic jetting. In this study, (AB)\(_{n}\) segmented copolymers are tailored for melt‐processing to form physically crosslinked hydrogels after swelling. The copolymers are composed of hydrophilic poly(ethylene glycol)‐based segments and hydrophobic bisurea segments, which form physical crosslinks via hydrogen bonds. The degree of polymerization was adjusted to match the melt viscosity to the different melt‐processing techniques. Using extrusion‐based printing, a width of approximately 260 µm is printed into 3D constructs, with excellent interlayer bonding at fiber junctions, due to hydrogen bonding between the layers. For melt electrospinning, much thinner fibers in the range of about 1–15 µm are obtained and produced in a typical nonwoven morphology. With melt electrowriting, fibers are deposited in a controlled way to well‐defined 3D constructs. In this case, multiple fiber layers fuse together enabling constructs with line width in the range of 70 to 160 µm. If exposed to water the printed constructs swell and form physically crosslinked hydrogels that slowly disintegrate, which is a feature for soluble inks within biofabrication strategies. In this context, cytotoxicity tests confirm the viability of cells and thus demonstrating biocompatibility of this class of copolymers. KW - 3D printing KW - (AB)\(_{n}\) segmented copolymers KW - biocompatibility KW - melt electrowriting Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-224513 VL - 222 IS - 1 ER - TY - JOUR A1 - Liashenko, Ievgenii A1 - Hrynevich, Andrei A1 - Dalton, Paul D. T1 - Designing Outside the Box: Unlocking the Geometric Freedom of Melt Electrowriting using Microscale Layer Shifting JF - Advanced Materials N2 - Melt electrowriting, a high‐resolution additive manufacturing technology, has so far been developed with vertical stacking of fiber layers, with a printing trajectory that is constant for each layer. In this work, microscale layer shifting is introduced through deliberately offsetting the printing trajectory for each printed layer. Inaccuracies during the printing of sinusoidal walls are corrected via layer shifting, resulting in accurate control of their geometry and mechanical properties. Furthermore, more substantial layer shifting allows stacking of fiber layers in a horizontal manner, overcoming the electrostatic autofocusing effect that favors vertical layer stacking. Novel nonlinear geometries, such as overhangs, wall texturing and branching, and smooth and abrupt changes in printing trajectory are presented, demonstrating the flexibility of the layer shifting approach beyond the state‐of‐the‐art. The practice of microscale layer shifting for melt electrowriting enables more complex geometries that promise to have a profound impact on the development of products in a broad range of applications. KW - 3D printing KW - additive manufacturing KW - biomaterials KW - electrohydrodynamics KW - melt electrospinning writing Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-217974 VL - 32 IS - 28 ER - TY - JOUR A1 - Kade, Juliane C. A1 - Tandon, Biranche A1 - Weichhold, Jan A1 - Pisignano, Dario A1 - Persano, Luana A1 - Luxenhofer, Robert A1 - Dalton, Paul D. T1 - Melt electrowriting of poly(vinylidene fluoride‐co‐trifluoroethylene) JF - Polymer International N2 - Poly(vinylidene fluoride-co-trifluoroethylene) (P(VDF-co-TrFE)) is an electroactive polymer with growing interest for applications in biomedical materials and flexible electronics. In this study, a solvent-free additive manufacturing technique called melt electrowriting (MEW) has been utilized to fabricate well-defined microperiodic structures of the copolymer (P(VDF-co-TrFE)). MEW of the highly viscous polymer melt was initiated using a heated collector at temperatures above 120 °C and required remarkably slow collector speeds below 100 mm min\(^{-1}\). The fiber surface morphology was affected by the collector speed and an increase in β-phase was observed for scaffolds compared to the unprocessed powder. Videography shows vibrations of the P(VDF-co-TrFE) jet previously unseen during MEW, probably due to repeated charge buildup and discharge. Furthermore, piezo-force microscopy measurements demonstrated the electromechanical response of MEW-fabricated fibers. This research therefore achieves the melt electrohydrodynamic processing of fibers with micrometer resolution into defined structures with an important electroactive polymer. KW - polymer processing KW - additive manufacturing KW - electrohydrodynamic KW - electroactive Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-257654 VL - 70 IS - 12 ER -