In vitro generation of colonic epithelium from primary cells guided by microstructures

Lab Chip. 2014 May 7;14(9):1622-31. doi: 10.1039/c3lc51353j. Epub 2014 Mar 20.


The proliferative compartment of the colonic epithelium in vivo is located in the basal crypt where colonic stem cells and transit-amplifying cells reside and fuel the rapid renewal of non-proliferative epithelial cells as they migrate toward the gut lumen. To mimic this tissue polarity, microstructures composed of polydimethylsiloxane (PDMS) microwells and Matrigel micropockets were used to guide a combined 2-dimensional (2D) and 3-dimensional (3D) hybrid culture of primary crypts isolated from the murine colon. The 2D and 3D culture of crypts on a planar PDMS surface was first investigated in terms of cell proliferation and stem cell activity. 3D culture of crypts with overlaid Matrigel generated enclosed, but highly proliferative spheroids (termed colonoids). 2D culture of crypts produced a spreading monolayer of cells, which were non-proliferative. A combined 2D/3D hybrid culture was generated in a PDMS microwell platform on which crypts were loaded by centrifugation into microwells (diameter = 150 μm, depth = 150 μm) followed by addition of Matrigel that formed micropockets locking the crypts within the microwells. Embedded crypts first underwent 3D expansion inside the wells. After the cells filled the microwells, they migrated onto the surrounding surface forming a 2D monolayer in the array regions without Matrigel. This unique 2D/3D hybrid culture generated a continuous, millimeter-scale colonic epithelial tissue in vitro, which resembled the polarized architecture (i.e. distinct proliferative and non-proliferative zones) and geometry of the colonic epithelium in vivo. This work initiates the construction of a "colon-on-a-chip" using primary cells/tissues with the ultimate goal of producing the physiologic structure and organ-level function of the colon.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Culture Techniques / methods*
  • Cell Proliferation
  • Cell Separation
  • Colon / cytology*
  • Dimethylpolysiloxanes / chemistry
  • Intestinal Mucosa / cytology*
  • Mice
  • Tissue Array Analysis / methods*


  • Dimethylpolysiloxanes
  • baysilon