title: The use of patterned dual thermoresponsive surfaces for the collective recovery as co-cultured cell sheets.
abstract: Heterotypic cell interactions are critical to achieve and maintain specific functions in many tissues and organs. We have focused on patterned structure surfaces to enable co-culture of heterotypic cells and recovery of patterned co-cultured cell sheets for applications in tissue engineering. Thermoresponsive polymers exhibiting different transition temperatures in water comprise both poly(N-isopropylacrylamide) (PIPAAm) and n-butyl methacrylate (BMA) co-grafted as side chains to PIPAAm main chains. These copolymers were surface-grafted in patterns to obtain patterned dual thermoresponsive cell culture surfaces using electron beam polymerisation method and porous metal masks. On patterned surfaces, site-selective adhesion on and growth of rat primary hepatocytes (HCs) and bovine carotid endothelial cells (ECs) allowed patterned co-culture, exploiting hydrophobic/hydrophilic surface chemistry regulated by culture temperature as the sole variable. At 27 degrees C, seeded HCs adhered exclusively onto hydrophobic, dehydrated P(IPAAm-BMA) co-grafted domains (1-mm laser dot), but not onto neighbouring hydrated PIPAAm domains. Sequentially seeded ECs then adhered exclusively to hydrophobised PIPAAm domains upon increasing culture temperature to 37 degrees C, achieving patterned co-cultures. Reducing culture temperature to 20 degrees C promoted hydration of both polymer-grafted domains, permitting release of the co-cultured, patterned cell monolayers as continuous cell sheets with heterotypic cell interactions. Recovered co-cultured cell sheets can be manipulated, moved and sandwiched with other structures, providing new useful constructs both for basic cell biology research and preparation of tissue-mimicking multi-layer materials through overlaying co-cultured cell sheets.
authors.affiliation: Graduate School of Science and Engineering, Waseda University, 3-4-1 Okubo, Shinjuku, Tokyo 169-8555, Japan.
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Tsuda
authors.forename: Akihiko
authors.initials: A
authors.lastname: Kikuchi
authors.forename: Masayuki
authors.initials: M
authors.lastname: Yamato
authors.forename: Aiko
authors.initials: A
authors.lastname: Nakao
authors.forename: Yasuhisa
authors.initials: Y
authors.lastname: Sakurai
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Umezu
authors.forename: Teruo
authors.initials: T
authors.lastname: Okano
mesh_terms: D000180:Acrylic Resins / Q000737:chemistry* / Q000528:radiation effects; D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry; D002417:Cattle; D002448:Cell Adhesion / Q000502:physiology; D002478:Cells, Cultured; D018920:Coculture Techniques / Q000379:methods*; D042783:Endothelial Cells / Q000166:cytology* / Q000502:physiology; D022781:Hepatocytes / Q000166:cytology* / Q000502:physiology; D006358:Hot Temperature; D008297:Male; D008422:Materials Testing; D008689:Methacrylates / Q000737:chemistry* / Q000528:radiation effects; D051381:Rats; D017208:Rats, Wistar; D013499:Surface Properties; D013696:Temperature; D023822:Tissue Engineering / Q000379:methods*
chemical_list: D000180:Acrylic Resins; D001672:Biocompatible Materials; D008689:Methacrylates; C052970:poly-N-isopropylacrylamide; C016284:butyl methacrylate