Recombinant laminin fragments endowed with collagen-binding activity: A tool for conferring laminin-like cell-adhesive activity to collagen matrices

Matrix Biol. 2018 Jan;65:75-90. doi: 10.1016/j.matbio.2017.08.001. Epub 2017 Aug 8.

Abstract

Laminins are major components of basement membranes that sustain a wide variety of stem cells. Among 15 laminin isoforms, laminin-511 and its E8 fragment (LM511E8) have been shown to strongly promote the adhesion and proliferation of human pluripotent stem cells. The aim of this study was to endow the cell-adhesive activity of laminin-511 on collagen matrices, thereby fabricating collagen-based culture scaffolds for stem cells with defined composition. To achieve this goal, we utilized the collagen-binding domain (CBD) of fibronectin to immobilize LM511E8 on collagen matrices. CBD was attached to the N-termini of individual laminin chains (α5E8, β1E8, γ1E8), producing LM511E8s having one, two, or three CBDs. While LM511E8 did not bind to collagen, CBD-attached LM511E8s (CBD-LM511E8s) exhibited significant collagen-binding activity, dependent on the number of attached CBDs. Human iPS cells were cultured on collagen-coated plates preloaded with CBD-LM511E8s. Although iPS cells did not attach or grow on collagen, they robustly proliferated on CBD-LM511E8-loaded collagen matrices, similar to the case with LM511E8-coated plates. Importantly, iPS cells proliferated and yielded round-shaped colonies even on collagen gels preloaded with CBD-LM511E8s. These results demonstrate that CBD-attached laminin E8 fragments are promising tools for fabrication of collagen-based matrices having the cell-adhesive activity of laminins.

Keywords: Collagen; Fibronectin; Laminin; iPS cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Basement Membrane / metabolism
  • Binding Sites
  • Cell Adhesion
  • Cell Culture Techniques
  • Cell Line
  • Cell Proliferation
  • Collagen / metabolism*
  • Fibronectins / chemistry*
  • Fibronectins / metabolism
  • Humans
  • Immobilized Proteins / metabolism
  • Induced Pluripotent Stem Cells / cytology*
  • Induced Pluripotent Stem Cells / metabolism
  • Laminin / chemistry*
  • Laminin / metabolism
  • Mice
  • NIH 3T3 Cells
  • Recombinant Proteins / metabolism*
  • Tissue Scaffolds

Substances

  • Fibronectins
  • Immobilized Proteins
  • Laminin
  • Recombinant Proteins
  • laminin-511, human
  • Collagen