title: Reconstruction of Highly Proliferative Auto-Tissue-Engineered Lamellar Cornea Enhanced by Embryonic Stem Cell.
abstract: To increase the epithelial proliferation of an auto-tissue-engineered lamellar cornea, 3.0 × 10(6) corneal epithelial cells (CECs) were combined with 3 × 10(5) mouse embryonic stem cells (ESCs) pretransfected with the HSV-tk gene (CECs + ESCs-TK group), and 3.3×10(6) corneal epithelial cells (CECs group) were seeded between the acellular porcine corneal stroma and the amniotic membrane using the centrifugal cell seeding method. After 4 days of perfusion culture (treatment with ganciclovir starting on day 2), a thicker corneal epithelium (four to five layers) formed in the CECs + ESCs-TK group compared with that observed in the CECs group (two to three layers). More stem/progenitor cell (K3 -, p63+, ABCG2+, and integrin-β1+) and proliferation phenotypes (Ki67+) were measured in the CECs + ESCs-TK group compared with the CECs group using immunofluorescence staining, real-time quantitative reverse transcription polymerase chain reaction, and flow cytometry. Consistent with these findings, the colony-forming efficiency and cellular doubling time were significantly different between the CECs + ESCs-TK group (16.18% ± 3.98%, 28.45 ± 2.03 h) and CECs group (11.96% ± 2.60%, 36.3 ± 1.15 h). In a rabbit lamellar transplantation model, the CECs + ESCs-TK group had better epithelial barrier functions and wound healing abilities compared with the CECs group. Furthermore, ESCs-TK could be completely and safely removed by ganciclovir. Thus, the ESCS-TK coculture system could serve as a potential strategy for corneal tissue engineering.
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Zhao
authors.initials: Z
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Zheng
authors.initials: Z
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Xiaoran
authors.initials: X
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Bowen
authors.initials: B
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Chaoyang
authors.initials: C
authors.lastname: Li
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Liangliang
authors.initials: L
authors.lastname: Li
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Pengxia
authors.initials: P
authors.lastname: Wan
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Zheqian
authors.initials: Z
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University , Guangzhou, China .
authors.forename: Zhichong
authors.initials: Z
authors.lastname: Wang
mesh_terms: D000818:Animals; D049109:Cell Proliferation; D018920:Coculture Techniques; D003315:Cornea / Q000166:cytology*; D053595:Embryonic Stem Cells / Q000166:cytology*; D005260:Female; D015774:Ganciclovir / Q000008:administration & dosage; D006801:Humans; D008297:Male; D051379:Mice; D011817:Rabbits; D013552:Swine; D023822:Tissue Engineering
chemical_list: D015774:Ganciclovir