Technique article: An experimental model for assessing fibroblast migration in 3-D collagen matrices

Dimitris Karamichos, Neema Lakshman, W. Matthew Petroll

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The purpose of this study was to develop and test a novel culture model for studying fibroblast migration in 3-D collagen matrices. Human corneal fibroblasts were seeded within dense, randomly oriented compressed collagen matrices. A 6 mm diameter button of this cell-seeded matrix was placed in the middle of an acellular, less dense outer collagen matrix. These constructs were cultured for 1, 3, 5 or 7 days in serum-free media, 10% fetal bovine serum (FBS), or 50 ng/ml PDGF. Constructs were then fixed and labeled with AlexaFluor 546 phalloidin (for f-actin) and TOTG-3 (for nuclei). Cell-matrix interactions were assessed using a combination of fluorescent and reflected light confocal imaging. Human corneal fibroblasts in serum-free media showed minimal migration into the outer (non-compressed) matrix. In contrast, culture in serum or PDGF stimulated cell migration. Cell-induced collagen matrix reorganization in the outer matrix could be directly visualized using reflected light imaging, and was highest following culture in 10% FBS. Cellular contraction in 10% FBS often led to alignment of cells parallel to the outer edge of the inner matrix, similar to the pattern observed during corneal wound healing following incisional surgery. Overall, this 3-D model allows the effects of different culture conditions on cell migration and matrix remodeling to be assessed simultaneously. In addition, the design allows for ECM density, geometry and mechanical constraints to be varied in a controlled fashion. These initial results demonstrate differences in cell and matrix patterning during migration in response to serum and PDGF.

Original languageEnglish (US)
Pages (from-to)1-9
Number of pages9
JournalCell Motility and the Cytoskeleton
Volume66
Issue number1
DOIs
StatePublished - Jan 2009

Fingerprint

Theoretical Models
Collagen
Fibroblasts
Serum
Serum-Free Culture Media
Cell Movement
Phalloidine
Light
Cell Communication
Wound Healing
Actins

Keywords

  • Cell migration
  • Confocal microscopy
  • Corneal fibroblasts
  • Extracellular matrix
  • PDGF

ASJC Scopus subject areas

  • Cell Biology
  • Structural Biology

Cite this

Technique article : An experimental model for assessing fibroblast migration in 3-D collagen matrices. / Karamichos, Dimitris; Lakshman, Neema; Petroll, W. Matthew.

In: Cell Motility and the Cytoskeleton, Vol. 66, No. 1, 01.2009, p. 1-9.

Research output: Contribution to journalArticle

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abstract = "The purpose of this study was to develop and test a novel culture model for studying fibroblast migration in 3-D collagen matrices. Human corneal fibroblasts were seeded within dense, randomly oriented compressed collagen matrices. A 6 mm diameter button of this cell-seeded matrix was placed in the middle of an acellular, less dense outer collagen matrix. These constructs were cultured for 1, 3, 5 or 7 days in serum-free media, 10{\%} fetal bovine serum (FBS), or 50 ng/ml PDGF. Constructs were then fixed and labeled with AlexaFluor 546 phalloidin (for f-actin) and TOTG-3 (for nuclei). Cell-matrix interactions were assessed using a combination of fluorescent and reflected light confocal imaging. Human corneal fibroblasts in serum-free media showed minimal migration into the outer (non-compressed) matrix. In contrast, culture in serum or PDGF stimulated cell migration. Cell-induced collagen matrix reorganization in the outer matrix could be directly visualized using reflected light imaging, and was highest following culture in 10{\%} FBS. Cellular contraction in 10{\%} FBS often led to alignment of cells parallel to the outer edge of the inner matrix, similar to the pattern observed during corneal wound healing following incisional surgery. Overall, this 3-D model allows the effects of different culture conditions on cell migration and matrix remodeling to be assessed simultaneously. In addition, the design allows for ECM density, geometry and mechanical constraints to be varied in a controlled fashion. These initial results demonstrate differences in cell and matrix patterning during migration in response to serum and PDGF.",
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