A dual-layer silk fibroin scaffold for reconstructing the human corneal limbus

Biomaterials. 2012 May;33(13):3529-38. doi: 10.1016/j.biomaterials.2012.01.045. Epub 2012 Feb 13.


Membranes prepared from Bombyx mori silk fibroin have shown potential as a substrate for human limbal epithelial (L-EC) and stromal cell cultivation. Here we present fibroin as a dual-layer construct containing both an epithelium and underlying stroma for corneolimbal reconstruction. We have compared the growth and phenotype of L-EC on non-porous versus porous fibroin membranes. Furthermore, we have compared the growth of limbal mesenchymal stromal cells (L-MSC) in either serum-supplemented medium or the MesenCult-XF(®) culture system within fibroin fibrous mats. The co-culture of L-EC and L-MSC in fibroin dual-layer constructs was also examined. L-EC on porous membranes displayed a squamous monolayer; in contrast, L-EC on non-porous fibroin appeared cuboidal and stratified. Both constructs maintained evidence of corneal phenotype (cytokeratin 3/12) and distribution of ΔNp63(+) progenitor cells. L-MSC cultivated within fibroin fibrous mats in serum-supplemented medium contained less than 64% of cells expressing the characteristic MSC phenotype of CD73(+)CD90(+)CD105(+) after two weeks, compared with over 81% in MesenCult-XF(®) medium. Dual-layer fibroin scaffolds consisting of L-EC and L-MSC maintained a similar phenotype as on the separate layers. These results support the feasibility of a 3D engineered limbus constructed from B. mori silk fibroin, and warrant further studies into the potential benefits it offers to corneolimbal tissue regeneration.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bombyx
  • Cell Adhesion / drug effects
  • Cell Proliferation / drug effects
  • Coculture Techniques
  • Epithelial Cells / cytology
  • Epithelial Cells / drug effects
  • Epithelial Cells / ultrastructure
  • Fibroins / metabolism*
  • Fibroins / pharmacology
  • Fibroins / ultrastructure
  • Humans
  • Immunophenotyping
  • Limbus Corneae / drug effects
  • Limbus Corneae / physiology*
  • Membranes, Artificial
  • Mesenchymal Stem Cells / cytology
  • Mesenchymal Stem Cells / drug effects
  • Mesenchymal Stem Cells / ultrastructure
  • Porosity / drug effects
  • Tissue Engineering / methods*
  • Tissue Scaffolds / chemistry*


  • Membranes, Artificial
  • Fibroins