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Bone-forming cells with pronounced spread into the third dimension in polymer scaffolds fabricated by two-photon polymerization.

ABSTRACT: The main aim of this work was to stimulate bone-forming cells to produce three-dimensional networks of mineralized proteins such as those occurring in bones. This was achieved by a novel approach using a specific type of mesenchymal progenitor cells (i.e., primary fibroblast cells from human hair roots) seeded on to polymer scaffolds. We wrote polymer microstructures with one or more levels of quadratic pores on to a flexible substrate by means of two-photon polymerization using a Ti-sapphire femtosecond laser focused into a liquid acrylate-based resin containing a photoinitiator. Progenitor cells, differentiated into an osteogenic lineage by the use of medium supplemented with biochemical stimuli, can be seeded on to the hydrophilic three-dimensional scaffolds. Due to confinement to the microstructures and/or mechanical interaction with the scaffold, the cells are stimulated to produce high amounts of calcium-binding proteins, such as collagen type I, and show an increased activation of the actin cytoskeleton. The best results were obtained for quadratic pore sizes of 35 µm: the pore volumes become almost filled with both cells in close contact with the walls of the structure and with extracellular matrix material produced by the cells. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 891-899, 2017.

SUBMITTER: Heitz J

PROVIDER: S-EPMC5299529 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Bone-forming cells with pronounced spread into the third dimension in polymer scaffolds fabricated by two-photon polymerization.

Heitz J J Plamadeala C C Wiesbauer M M Freudenthaler P P Wollhofen R R Jacak J J Klar T A TA Magnus B B Köstner D D Weth A A Baumgartner W W Marksteiner R R

Journal of biomedical materials research. Part A 20161205 3

The main aim of this work was to stimulate bone-forming cells to produce three-dimensional networks of mineralized proteins such as those occurring in bones. This was achieved by a novel approach using a specific type of mesenchymal progenitor cells (i.e., primary fibroblast cells from human hair roots) seeded on to polymer scaffolds. We wrote polymer microstructures with one or more levels of quadratic pores on to a flexible substrate by means of two-photon polymerization using a Ti-sapphire fe ...[more]

PMID: 27813317

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Project description:An understanding of parameters that modulate gene transfer in 3-D will assist in the formation of gene delivery systems and scaffolds, which can mediate efficient non-viral delivery for guiding in vivo tissue regeneration and therapy. We have previously demonstrated the cell area and length, integrin expression, and RhoGTPase mediated signalling to be pivotal parameters that guide gene transfer to mouse mesenchymal stem cells (mMSCs) cultured in 2-D and are modulated by ECM proteins. In this study, we were interested in determining if cationic polymer mediated gene transfer to cells seeded in 3-D would occur through different mechanisms as compared to those seeded in 2-D. In particular, we examined the endocytosis pathways used to internalize polyplexes, and the role of cytoskeletal dynamics and RhoGTPases in non-viral gene transfer for cells seeded in 2-D and 3-D. Inhibition of clathrin- and caveolae-mediated endocytosis resulted in a more drastic decrease in overall transgene expression for cells seeded in 3-D than for those in 2-D. In addition, polyplex internalization was only significantly decreased in 3-D when clathrin-mediated endocytosis was inhibited, while caveolae-mediated endocytosis inhibition for cells seeded in 2-D resulted in the strongest polyplex internalization inhibition. Actin and microtubule polymerization affected 2-D and 3-D transfection differently. Microtubule depolymerization enhanced transgene expression in 2-D, but inhibited transgene expression in 3-D. Lastly, inhibition of RhoGTPases also affected 2-D and 3-D transfection differently. The inhibition of ROCK effectors resulted in a decrease of transgene expression and internalization for cells seeded in 3-D, but not in 2-D, and the inhibition of the effector PAK1 resulted in an increase of transgene expression for both 2-D and 3-D. Overall, our study suggests that the process of gene transfer occurs through different mechanisms for cells seeded in 2-D compared to those seeded in 3-D.

Dataset Information

Bone-forming cells with pronounced spread into the third dimension in polymer scaffolds fabricated by two-photon polymerization.

Publications

Bone-forming cells with pronounced spread into the third dimension in polymer scaffolds fabricated by two-photon polymerization.

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