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Author Nithya, Joseph ♦ Kumar, P. R. ♦ Tilak, Prasad ♦ Leena, Joseph ♦ Sreenivasan, K. ♦ Kumari, T. V.
Source Sree Chitra Tirunal Institute for Medical Sciences & Technology
Content type Text
Publisher Tissue Engineering Part C-methods
File Format PDF
Language English
Subject Domain (in DDC) Technology ♦ Medicine & health ♦ Human physiology
Subject Domain (in MeSH) Tissues ♦ Cells ♦ Anatomy ♦ Investigative Techniques ♦ Analytical, Diagnostic and Therapeutic Techniques and Equipment
Subject Keyword Tissue Engineering
Abstract Cell sheet engineering using thermoresponsive culture dishes allows harvesting of intact in vitro cell sheet. In this study, commercially available polyethylene terephthalate-based overhead projection transparency sheet (OHPS) was identified as a substrate for coating thermoresponsive poly(N-isopropylacrylamide-co-glycidylmethacrylate) (NGMA) copolymer having lower critical solution temperature of 28 degrees C. Since OHPS is highly hydrophobic and rigid, the surface was modified by alkali treatment (OHPS-M) to functionalize the surface with carboxyl and hydroxyl groups so as to make it more suitable for efficient coating of NGMA copolymer and cell culture. To impart thermoresponsiveness, OHPS-M was coated with NGMA (OHPS-MC). Surface morphology, surface chemistry, and thermoresponsive coating were analyzed by profilometry, scanning electron microscopy, water contact angle, and Fourier transform infrared spectroscopy. The cytotoxicity, cell adhesion, and proliferation on OHPS-M and OHPS-MC were analyzed using L929 cells. Specific cytocompatibility analysis was done using SIRC (Rabbit corneal) cells. Data revealed cytocompatible nature of OHPS-M and OHPS-MC. Suitability of OHPS-MC for cell sheet harvest and transfer efficiency was assessed using primary corneal cells. Corneal cell sheet constructs retrieved by temperature variation was characterized by reverse transcriptase polymerase chain reaction for markers specific to differentiated corneal cells (keratin 3 and keratin 12) and proliferating cell nuclear antigen, and assessed for viability using fluorescein diacetate staining, tissue architecture by scanning electron microscopy, and cell-cell contacts by connexin-43 staining. The retrieved cell sheets retained corneal epithelial characteristics such as keratin 3/12, viability, and tissue architecture with intact cell-cell contacts as in native tissue. The results proved that surface modification and coating with NGMA on OHPS offer a novel biocompatible thermosensitive cell culture substrate for generating cell sheet constructs. In addition, OHPS-M can also serve as an efficient carrier tool for retrieved cell sheet.
Education Level UG and PG
Learning Resource Type Article
Educational Framework Medical Council of India (MCI)
Volume Number 17
Issue Number 2
Page Count 11
Starting Page 181
Ending Page 191