Culture and differentiation of rat neural stem/progenitor cells in a three-dimensional collagen scaffold

Appl Biochem Biotechnol. 2013 May;170(2):406-19. doi: 10.1007/s12010-013-0211-5. Epub 2013 Mar 28.

Abstract

A stable and fast method for constructing a neural-like tissue from rat neural stem/progenitor cells (rNS/PCs) based on three-dimensional (3D) collagen gel is described. First step, the collagen-embedded rNS/PCs expanded with the medium consisting of DMEM/F12/RPMI1640 (1:1:1) supplemented with EGF and bFGF was used to expand the cells in gel in 96-well plates until the average diameter of cell clusters was about 50-100 μm with the cell density higher than 10(7) cells/mL. In the second step, the initial medium was replaced with NB/B-27 supplemented with bFGF and BDNF. The results show that cells in collagen presented neural-like morphology and maintained live cell rate around 82 % in neural network pattern at least for 42 days under static conditions. The cell-collagen constructs were detected by immunofluorescence and immunohistochemistry test after 42 days of culture, part of cells still maintained the character of rNS/PCs, and others differentiated into neurons, astrocytes, and oligodendrocytes. Our 3D neural-like tissue construct was similar to the neural tissue in morphology and cell compositions. They thus have a potential to be used for drug screening, detection of environment toxins, and replacement therapy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Astrocytes / cytology
  • Astrocytes / metabolism
  • Brain-Derived Neurotrophic Factor / pharmacology
  • Cell Count
  • Cell Culture Techniques
  • Cell Differentiation*
  • Cell Proliferation
  • Cell Shape*
  • Cell Survival
  • Cells, Cultured
  • Collagen / metabolism*
  • Culture Media / metabolism
  • Fibroblast Growth Factor 2 / pharmacology
  • Hippocampus / cytology
  • Hippocampus / metabolism
  • Imaging, Three-Dimensional
  • Neural Stem Cells / cytology*
  • Neural Stem Cells / metabolism
  • Neurons / cytology
  • Neurons / metabolism
  • Rats
  • Rats, Sprague-Dawley
  • Tissue Scaffolds*

Substances

  • Brain-Derived Neurotrophic Factor
  • Culture Media
  • Fibroblast Growth Factor 2
  • Collagen