title: Acellular porcine corneal matrix as a carrier scaffold for cultivating human corneal epithelial cells and fibroblasts in vitro.
abstract: OBJECTIVE
  To investigate the feasibility of corneal anterior lamellar reconstruction with human corneal epithelial cells and fibroblasts, and an acellular porcine cornea matrix (APCM) in vitro.
  
  
  METHODS
  The scaffold was prepared from fresh porcine corneas which were treated with 0.5% sodium dodecyl sulfate (SDS) solution and the complete removal of corneal cells was confirmed by hematoxylin-eosin (HE) staining and 4', 6-diamidino-2-phenylindole (DAPI) staining. Human corneal fibroblasts and epithelial cells were cultured with leaching liquid extracted from APCM, and then cell proliferative ability was evaluated by MTT assay. To construct a human corneal anterior lamellar replacement, corneal fibroblasts were injected into the APCM and cultured for 3d, followed by culturing corneal epithelial cells on the stroma construction surface for another 10d. The corneal replacement was analyzed by HE staining, and immunofluorescence staining.
  
  
  RESULTS
  Histological examination indicated that there were no cells in the APCM by HE staining, and DAPI staining did not detect any residual DNA. The leaching liquid from APCM had little influence on the proliferation ability of human corneal fibroblasts and epithelial cells. At 10d, a continuous 3 to 5 layers of human corneal epithelial cells covering the surface of the APCM was observed, and the injected corneal fibroblasts distributed within the scaffold. The phenotype of the construction was similar to normal human corneas, with high expression of cytokeratin 12 in the epithelial cell layer and high expression of vimentin in the stroma.
  
  
  CONCLUSIONS
  Corneal anterior lamellar replacement can be reconstructed in vitro by cultivating human corneal epithelial cells and fibroblasts with an acellular porcine cornea matrix. This laid the foundation for the further transplantation in vivo.
authors.affiliation: Department of Ophthalmology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China; The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Health, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China.
authors.forename: Ju
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Ophthalmology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China.
authors.forename: Can-Wei
authors.initials: CW
authors.lastname: Zhang
authors.affiliation: Department of Ophthalmology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China.
authors.forename: Li-Qun
authors.initials: LQ
authors.lastname: Du
authors.affiliation: Department of Ophthalmology, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China.
authors.forename: Xin-Yi
authors.initials: XY
authors.lastname: Wu
keywords: acellular porcine cornea matrix; corneal epithelial cells; corneal keratocytes; corneal tissue engineering; limbal epithelial cells