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An Innovative Strategy for the Fabrication of Functional Cell Sheets Using an Electroactive Conducting Polymer.

Lee H, Cho Y - Theranostics (2015)

Bottom Line: Here, we report the development of an electric field-assisted methodology for constructing 3D C2C12 cell sheets with the potential for cell surface modification.Subsequently, C2C12 cells are cultured on BMP2-immobilized Ppy surfaces to induce interactions between cell surface receptors and bound BMP2 ligands.Following these procedures, layers of BMP2-immobilized cells can be easily detached from the Ppy surface by applying an electrical potential.

View Article: PubMed Central - PubMed

Affiliation: New Experimental Therapeutic Branch, National Cancer Center, 111 Jungbalsan-ro, Ilsandong-gu, Goyang, Gyeonggi-do 410-769, South Korea.

ABSTRACT
Here, we report the development of an electric field-assisted methodology for constructing 3D C2C12 cell sheets with the potential for cell surface modification. In this method, a conducting polymer, polypyrrole (Ppy), is electrodeposited via biotin doping, and then chemical conjugation of biotinylated bone morphogenetic protein 2 (BMP2) is achieved using a biotin-streptavidin cross-linker. Subsequently, C2C12 cells are cultured on BMP2-immobilized Ppy surfaces to induce interactions between cell surface receptors and bound BMP2 ligands. Following these procedures, layers of BMP2-immobilized cells can be easily detached from the Ppy surface by applying an electrical potential. This novel method results in high affinity, ligand-bound cell sheets, which exhibit homogeneous coverage with membrane-bound proteins and signal activation that occurs via maximal receptor accessibility. Using this strategy to engineer the cell surface with desirable ligands results in structures that mimic in vivo tissues; thus, the method reported here has potential applications in regenerative medicine and tissue engineering.

No MeSH data available.


Schematic diagram of the fabrication process for 3D cell sheets specifically conjugated with bone morphogenetic protein 2 (BMP2). C2C12 cells were incubated on BMP2-immobilized, biotin-doped polypyrrole (Ppy) surfaces. Cell sheets were produced with individual cells efficiently tethered to growth factors (specifically BMP2) via cell surface receptors, and these could be non-destructively released from Ppy by applying an electric field (-0.8 V for 30 s). Recovered cell sheets could be made to overlap as a 3D-multilayer.
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Figure 1: Schematic diagram of the fabrication process for 3D cell sheets specifically conjugated with bone morphogenetic protein 2 (BMP2). C2C12 cells were incubated on BMP2-immobilized, biotin-doped polypyrrole (Ppy) surfaces. Cell sheets were produced with individual cells efficiently tethered to growth factors (specifically BMP2) via cell surface receptors, and these could be non-destructively released from Ppy by applying an electric field (-0.8 V for 30 s). Recovered cell sheets could be made to overlap as a 3D-multilayer.

Mentions: A schematic diagram of the fabrication process for the 3D cell sheets is shown in Figure 1. Initially, Ppy was electrochemically polymerized on an ITO surface by using biotin as a co-dopant in the Ppy film. Biotin can be employed as a bridge in conjugation with target biomolecules. With this approach, it was possible to fabricate electric-field-assisted cell sheets, in which the mouse skeletal muscle-derived C2C12 cell line was used as a working model. Importantly, individual cells within the 3D constructs could be efficiently tethered with growth factors, specifically bone morphogenetic protein 2 (BMP2), via cell surface receptors. BMP2 plays an important role in inducing osteoblastic differentiation of the C2C12 myoblasts by blocking the myogenic differentiation pathway.21, 22 The introduction of BMP2 in the vicinity of the cell surface increases the recognition of, as well as communication with, cell membrane receptors, which facilitates the stable formation of complexes between growth factors and receptors with sustained receptor activation. This strategy allows for the manipulation of individual target cells with desired functional entities and, subsequently, the modulation of cellular activity. Indeed, the guided assembly of biotin as a dopant is expected to achieve the homogeneous and spatial arrangement of target moieties. The advantages of the Ppy-based scaffold-free cell sheets produced in this study are as follows: (i) the spontaneous and reversible red-ox reaction of the biotin-doped Ppy surface enables controlled incorporation and release of biomolecules; (ii) successful binding of BMP2 with receptors on the extracellular side of the cellular membrane significantly enhances the osteogenic differentiation of C2C12 cells, and does so by efficient and site-specific delivery of functional proteins; (iii) the scaffold-free technology promotes improved integration with the surrounding tissues, mimicry of tissue function, and high cell density that together lead to more complete regeneration of tissue. The simplicity of its fabrication should allow our scaffold-free cell sheet to be integrated with in vitro tissue/organ models and be used in vivo cell-based therapy.


An Innovative Strategy for the Fabrication of Functional Cell Sheets Using an Electroactive Conducting Polymer.

Lee H, Cho Y - Theranostics (2015)

Schematic diagram of the fabrication process for 3D cell sheets specifically conjugated with bone morphogenetic protein 2 (BMP2). C2C12 cells were incubated on BMP2-immobilized, biotin-doped polypyrrole (Ppy) surfaces. Cell sheets were produced with individual cells efficiently tethered to growth factors (specifically BMP2) via cell surface receptors, and these could be non-destructively released from Ppy by applying an electric field (-0.8 V for 30 s). Recovered cell sheets could be made to overlap as a 3D-multilayer.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4493539&req=5

Figure 1: Schematic diagram of the fabrication process for 3D cell sheets specifically conjugated with bone morphogenetic protein 2 (BMP2). C2C12 cells were incubated on BMP2-immobilized, biotin-doped polypyrrole (Ppy) surfaces. Cell sheets were produced with individual cells efficiently tethered to growth factors (specifically BMP2) via cell surface receptors, and these could be non-destructively released from Ppy by applying an electric field (-0.8 V for 30 s). Recovered cell sheets could be made to overlap as a 3D-multilayer.
Mentions: A schematic diagram of the fabrication process for the 3D cell sheets is shown in Figure 1. Initially, Ppy was electrochemically polymerized on an ITO surface by using biotin as a co-dopant in the Ppy film. Biotin can be employed as a bridge in conjugation with target biomolecules. With this approach, it was possible to fabricate electric-field-assisted cell sheets, in which the mouse skeletal muscle-derived C2C12 cell line was used as a working model. Importantly, individual cells within the 3D constructs could be efficiently tethered with growth factors, specifically bone morphogenetic protein 2 (BMP2), via cell surface receptors. BMP2 plays an important role in inducing osteoblastic differentiation of the C2C12 myoblasts by blocking the myogenic differentiation pathway.21, 22 The introduction of BMP2 in the vicinity of the cell surface increases the recognition of, as well as communication with, cell membrane receptors, which facilitates the stable formation of complexes between growth factors and receptors with sustained receptor activation. This strategy allows for the manipulation of individual target cells with desired functional entities and, subsequently, the modulation of cellular activity. Indeed, the guided assembly of biotin as a dopant is expected to achieve the homogeneous and spatial arrangement of target moieties. The advantages of the Ppy-based scaffold-free cell sheets produced in this study are as follows: (i) the spontaneous and reversible red-ox reaction of the biotin-doped Ppy surface enables controlled incorporation and release of biomolecules; (ii) successful binding of BMP2 with receptors on the extracellular side of the cellular membrane significantly enhances the osteogenic differentiation of C2C12 cells, and does so by efficient and site-specific delivery of functional proteins; (iii) the scaffold-free technology promotes improved integration with the surrounding tissues, mimicry of tissue function, and high cell density that together lead to more complete regeneration of tissue. The simplicity of its fabrication should allow our scaffold-free cell sheet to be integrated with in vitro tissue/organ models and be used in vivo cell-based therapy.

Bottom Line: Here, we report the development of an electric field-assisted methodology for constructing 3D C2C12 cell sheets with the potential for cell surface modification.Subsequently, C2C12 cells are cultured on BMP2-immobilized Ppy surfaces to induce interactions between cell surface receptors and bound BMP2 ligands.Following these procedures, layers of BMP2-immobilized cells can be easily detached from the Ppy surface by applying an electrical potential.

View Article: PubMed Central - PubMed

Affiliation: New Experimental Therapeutic Branch, National Cancer Center, 111 Jungbalsan-ro, Ilsandong-gu, Goyang, Gyeonggi-do 410-769, South Korea.

ABSTRACT
Here, we report the development of an electric field-assisted methodology for constructing 3D C2C12 cell sheets with the potential for cell surface modification. In this method, a conducting polymer, polypyrrole (Ppy), is electrodeposited via biotin doping, and then chemical conjugation of biotinylated bone morphogenetic protein 2 (BMP2) is achieved using a biotin-streptavidin cross-linker. Subsequently, C2C12 cells are cultured on BMP2-immobilized Ppy surfaces to induce interactions between cell surface receptors and bound BMP2 ligands. Following these procedures, layers of BMP2-immobilized cells can be easily detached from the Ppy surface by applying an electrical potential. This novel method results in high affinity, ligand-bound cell sheets, which exhibit homogeneous coverage with membrane-bound proteins and signal activation that occurs via maximal receptor accessibility. Using this strategy to engineer the cell surface with desirable ligands results in structures that mimic in vivo tissues; thus, the method reported here has potential applications in regenerative medicine and tissue engineering.

No MeSH data available.