title: Autologous method for ex vivo expansion of human limbal epithelial progenitor cells based on plasma rich in growth factors technology.
abstract: Develop an autologous culture method for ex vivo expansion of human limbal epithelial progenitor cells (LEPCs) using Plasma Rich in Growth Factors (PRGF) as a growth supplement and as a scaffold for the culture of LEPCs.
  
  
  
  LEPCs were cultivated in different media supplemented with 10% fetal bovine serum (FBS) or 10% PRGF. The outgrowths, total number of cells, colony forming efficiency (CFE), morphology and immunocytochemistry against p63- α and cytokeratins 3 and 12 (CK3-CK12) were analyzed. PRGF was also used to elaborate a fibrin membrane. The effects of the scaffold on the preservation of stemness and the phenotypic characterization of LEPCs were investigated through analysis of CK3-CK12, ABCG-2 and p63.
  
  
  
  LEPCs cultivated with PRGF showed a significantly higher growth area than FBS cultures. Moreover, the number of cells were also higher in PRGF than FBS, while displaying a better morphology overall. CFE was found to be also higher in PRGF groups compared to FBS, and the p63-α expression also differed between groups. LEPCs cultivated on PRGF membranes appeared as a confluent monolayer of cells and still retained p63 and ABCG-2 expression, being negative for CK3-CK12.
  
  
  
  PRGF can be used in corneal tissue engineering, supplementing the culture media, even in a basal media without any other additives, as well as providing a scaffold for the culture.
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain. Electronic address: ana.riestra@fio.as.
authors.forename: A C
authors.initials: AC
authors.lastname: Riestra
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain.
authors.forename: N
authors.initials: N
authors.lastname: Vazquez
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain.
authors.forename: M
authors.initials: M
authors.lastname: Chacon
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain.
authors.forename: S
authors.initials: S
authors.lastname: Berisa
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain.
authors.forename: R M
authors.initials: RM
authors.lastname: Sanchez-Avila
authors.affiliation: Laboratory of Pharmacy and Pharmaceutical Technology, Faculty of Pharmacy, University of the Basque Country, Vitoria, Spain; Networking Biomedical Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, SLFPB-EHU, 01006 Vitoria-Gasteiz, Spain; BTI Biotechnology Institute, Vitoria, Spain.
authors.forename: G
authors.initials: G
authors.lastname: Orive
authors.affiliation: BTI Biotechnology Institute, Vitoria, Spain.
authors.forename: E
authors.initials: E
authors.lastname: Anitua
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain; U714 Centre for Biomedical Network Research on Rare Diseases (CIBERER), Oviedo, Spain.
authors.forename: A
authors.initials: A
authors.lastname: Meana
authors.affiliation: Instituto Oftalmológico Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Merayo-Lloves
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D003315:Cornea; D004847:Epithelial Cells; D006801:Humans; D016850:Limbus Corneae; D013234:Stem Cells
keywords: Cell culture techniques; Cell proliferation; Cell transplantation; Corneal epithelium; Culture media; Limbal deficiency; Limbus corneae; PRGF; Plasma rich in growth factors