title: Differentiation of rabbit bone marrow mesenchymal stem cells into corneal epithelial cells in vivo and ex vivo.
abstract: OBJECTIVE
  To examine whether bone marrow mesenchymal stem cells (MSCs) could be differentiated into corneal epithelial cells in vivo and ex vivo.
  
  
  METHODS
  In vivo, BrdU labeled rabbit MSCs (Rb-MSCs) were suspended in the fibrin gels and transplanted onto the surface of the damaged rabbit corneas. Histology and molecular phenotype were studied on postoperative day 28. In vitro, labeled Rb-MSCs were cultured for three days in two different systems: (1) Group A: Rb-MSCs were co-cultured with rabbit limbal stem cells (Rb-LSCs) by the Transwell culture system. A suspension of Rb-LSCs was added to the upper membrane surface, and the inserts were positioned in the culture wells, which were incubated with Rb-MSCs; (2) Group B: Supernatant medium that had first been used to culture Rb-LSCs and then filtered with a 0.45 mum filter was used to culture Rb-MSCs. For both groups, immunofluorescence and flow cytometric analysis were used to examine the expression of cytokeratin 3 (CK3) in differentiated Rb-MSCs.
  
  
  RESULTS
  In vivo, the data showed that following transplantation of Rb-MSCs, the rabbit's damaged corneal surface was successfully reconstructed and that some Rb-MSCs participated in the healing of the injured corneal epithelium and expressed CK3. In vitro, the data showed that Rb-MSCs rapidly differentiated into cells with a morphological and molecular phenotype of corneal epithelial-like cells. For both groups, the differentiated Rb-MSCs were positive for corneal epithelial-specific marker CK3. In Group A, flow cytometry analysis showed that at day one, only 3.46+/-1.9% of cells expressed CK3. This increased to 7.24+/-3.80% at day two and decreased slightly (5.50+/-3.33%) at day three. The proportion of CK3 in Group B was 4.09+/-1.84% at day one, rising to 9.31+/-5.92% after 24 h, but falling (4.37+/-2.61%) at day three. The mean differences are significant between each group and the negative control, but was not significant between Group A and Group B.
  
  
  CONCLUSIONS
  MSCs could differentiate into corneal epithelial-like cells in vivo and ex vivo.
authors.affiliation: China Medical University, Department of Ophthalmology, The First Affiliated Hospital, Shenyang, China.
authors.forename: Shaofeng
authors.initials: S
authors.lastname: Gu
authors.forename: Chengzhong
authors.initials: C
authors.lastname: Xing
authors.forename: Jingyi
authors.initials: J
authors.lastname: Han
authors.forename: Mark O M
authors.initials: MO
authors.lastname: Tso
authors.forename: Jing
authors.initials: J
authors.lastname: Hong
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology*; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D048430:Cell Shape; D004195:Disease Models, Animal; D019573:Epithelium, Corneal / Q000166:cytology*; D005337:Fibrin / Q000378:metabolism; D005434:Flow Cytometry; D016850:Limbus Corneae / Q000473:pathology; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D010641:Phenotype; D011817:Rabbits
chemical_list: D005337:Fibrin