Cell sheet patterning using photo-cleavable polymers

Abstract

We have found that poly (4-vinylpyridine) (P 4 VP) and poly (methylmethacrylate) thin films can be etched with ultraviolet A (UVA) radiation. Furthermore, we also found that dermal fibroblasts could be cultured successfully on the P 4 VP polymer, with a doubling time comparable to tissue culture Petri dish standards. Consequently, we were able to grow tissue on P 4 VP substrates, this could easily be lifted using UVA radiation. The cell sheets that were removed were then re-plated at a lower density and a series of assays was performed at 3 and 6 days. Although only a small amount of damage was discernable at day 3 nearly complete recovery was observed at day 6. The technique was used to pattern areas within the tissue, where other types of cells could be inserted. To demonstrate the technique, a hybrid tissue layer was produced, in which the dermal fibroblasts in a circular area at the center of the sample were removed by exposure through a mask. A keratinocyte layer was inserted, which adhere to the fibroblast layer forming a tissue with integrated layers of two distinct cell types. Polymer Journal (2011) 43, 723-732; doi:10.1038/pj.2011; published online 8 June 2011Keywords: biopolymers; cells INTRODUCTION A significant shortcoming of conventional tissue engineering methods is the loss of differentiated cell functions during cell collecting. Typical cell collecting techniques use proteolytic enzymes such as trypsin or dispase, which irreversibly damage cell surface proteins, such as growth factor receptors, ion channels and cell-to-cell junction proteins. 1 This result in an inability to assemble constructs from multiple cell types, which is an important requirement in tissue engineering.To address these shortcomings, cell sheet engineering has been proposed as a new methodology to facilitate the non-invasive collect of cultured cells as intact cell sheets. 2 As the cell sheets collected through this technique are recovered with intact cell-to-cell junctions and deposited extracellular matrix whole-cell sheets can be transferred to other tissue surfaces to produce versatile two-dimensional or threedimensional tissues. 3 Cell sheet engineering advanced rapidly with the development of culture dishes covalently grafted with temperatureresponsive polymers. These culture surfaces allowed whole, contiguous cell sheets to spontaneously detach from the surface simply by briefly decreasing the temperature. 4,5 This technique was also used to culture four layers of cardiomyocytes, which successfully formed pulsatile myocardial tissue in vitro and in vivo. 6 Other studies using epidermal keratinocytes have shown that it was possible to maintain the differentiated functions after the cell sheets were collected. 7 However, although thermoresponsive culture surfaces have shown success in producing whole sheets of hard, cell-dense tissues such as myocardium, the technique is not suitable for tissue patterning and for soft tissue. For example, biological soft tissue like brain and adipose are difficult to manipulate (for the...

Dynamism of Stimuli-Responsive Nanohybrids: Environmental Implications

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Effect of Graphene on Differentiation and Mineralization of Dental Pulp Stem Cells in Poly(4-vinylpyridine) Matrix in Vitro

“…In addition, P4VP does not require surface treatments and has better uniformity than tissue culture plastic of surface properties. 16 The results indicated that differentiation was determined by the ability of the collagen fibrils formed by the cells to cluster together and form the classical banded collagen fibers. Spuncast flat films and electrospun fiber scaffolds were compared, and banded collagen seemed to form only on the electrospun fibrous substrates.…”

“…Poly­(4-vinylpyridine) (P4VP) is a transparent polymer, which is glassy at ambient temperatures (glass transition, T g = 137 °C), and which has been shown to be equivalent to tissue culture plastic for cell culture, supporting adhesion, and proliferation of multiple cells types. In addition, P4VP does not require surface treatments and has better uniformity than tissue culture plastic of surface properties . The results indicated that differentiation was determined by the ability of the collagen fibrils formed by the cells to cluster together and form the classical banded collagen fibers.…”

Cell Patterning Technology on Polymethyl Methacrylate through Controlled Physicochemical and Biochemical Functionalization

“…In order to address PMMA's limitations, a variety of methods have been investigated in order to enhance cell adhesion on the material by increasing its surface hydrophilicity, including but not limited to chemical-and light-based treatments of the polymer [14][15][16][17]. However, one of the most facile and quick techniques to augment the surface hydrophilicity of a polymer is via air or oxygen plasma treatment.…”