title: Reconstruction of conjunctival epithelium-like tissue using a temperature-responsive culture dish.
abstract: OBJECTIVE
  To study the feasibility of engineering conjunctival epithelial cell sheets on a temperature-responsive culture dish for ocular surface reconstruction.
  
  
  METHODS
  Rabbit conjunctival epithelial cells (rCjECs) were cultured in DMEM/F-12 (1:1) medium. The morphology and phenotype of the rCjECs were confirmed with phalloidin staining, periodic acid-Schiff (PAS) staining, and immunocytochemistry. The rCjECs cultured on a temperature-responsive culture dish for 10 days produced confluent conjunctival epithelial cell sheets. Then, the phenotype, structure, and function of the conjunctival epithelial cell sheets were examined.
  
  
  RESULTS
  The conjunctival epithelial cells were compact, uniform, and cobblestone shape. All cultured conjunctival epithelial cells were harvested as intact cell sheets by reducing the culture temperature to 20 °C. Conjunctival epithelial cells were stratified in four to five cell layers similar to the conjunctival epithelium. CCK-8 analysis, 5-bromo-2'-deoxyuridine (BrdU) staining, and the live and dead viability assay confirmed that viable proliferation cells were retained in the cell sheets. Immunohistochemistry for CK4, CK19, and MUC5AC showed the cell sheets still maintained characteristics of the conjunctival epithelium.
  
  
  CONCLUSIONS
  A temperature-responsive culture dish enables fabrication of viable conjunctival epithelial cell sheets with goblet cells and proliferative cells. Conjunctival epithelial cell sheets will be promising for reconstruction of the conjunctival epithelium.
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Qinke
authors.initials: Q
authors.lastname: Yao
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Mengyu
authors.initials: M
authors.lastname: Zhu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Junzhao
authors.initials: J
authors.lastname: Chen
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Chunyi
authors.initials: C
authors.lastname: Shao
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Chenxi
authors.initials: C
authors.lastname: Yan
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Zi
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Xianqun
authors.initials: X
authors.lastname: Fan
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Ping
authors.initials: P
authors.lastname: Gu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
authors.forename: Yao
authors.initials: Y
authors.lastname: Fu
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003228:Conjunctiva / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D015870:Gene Expression; D053527:Keratin-4 / Q000235:genetics / Q000378:metabolism; D055271:Mucin 5AC / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D013696:Temperature; D023822:Tissue Engineering / Q000379:methods*
chemical_list: D015415:Biomarkers; D003470:Culture Media; D053527:Keratin-4; D055271:Mucin 5AC