Quantitative assessment of local collagen matrix remodeling in 3-D Culture: The role of Rho kinase

Areum Kim, Neema Lakshman, W. Matthew Petroll

Research output: Contribution to journalArticle

86 Citations (Scopus)

Abstract

The purpose of this study was to quantitatively assess the role of Rho kinase in modulating the pattern and amount of local cell-induced collagen matrix remodeling. Human corneal fibroblasts were plated inside 100-μm thick fibrillar collagen matrices and cultured for 24 h in media with or without the Rho kinase inhibitor Y-27632. Cells were then fixed and stained with phalloidin. Fluorescent (for f-actin) and reflected light (for collagen fibrils) 3-D optical section images were acquired using laser confocal microscopy. Fourier transform analysis was used to assess collagen fibril alignment, and 3-D cell morphology and local collagen density were measured using MetaMorph. Culture in serum-containing media induced significant global matrix contraction, which was inhibited by blocking Rho kinase (p < 0.001). Fibroblasts generally had a bipolar morphology and intracellular stress fibers. Collagen fibrils were compacted and aligned parallel to stress fibers and pseudopodia. When Rho kinase was inhibited, cells had a more cortical f-actin distribution and dendritic morphology. Both local collagen fibril density and alignment were significantly reduced (p < 0.01). Overall, the data suggests that Rho kinase-dependent contractile force generation leads to co-alignment of cells and collagen fibrils along the plane of greatest resistance, and that this process contributes to global matrix contraction.

Original languageEnglish (US)
Pages (from-to)3683-3692
Number of pages10
JournalExperimental Cell Research
Volume312
Issue number18
DOIs
StatePublished - Nov 1 2006

Fingerprint

rho-Associated Kinases
Collagen
Stress Fibers
Fourier Analysis
Confocal Microscopy
Actins
Fibroblasts
Fibrillar Collagens
Phalloidine
Pseudopodia
Somatostatin-Secreting Cells
Light
Serum

Keywords

  • Actin
  • Cell mechanics
  • Collagen matrices
  • Confocal microscopy
  • Corneal fibroblasts
  • Cytoskeleton
  • Fourier transform
  • Myosin
  • Rho kinase

ASJC Scopus subject areas

  • Cell Biology

Cite this

Quantitative assessment of local collagen matrix remodeling in 3-D Culture : The role of Rho kinase. / Kim, Areum; Lakshman, Neema; Petroll, W. Matthew.

In: Experimental Cell Research, Vol. 312, No. 18, 01.11.2006, p. 3683-3692.

Research output: Contribution to journalArticle

@article{59ea9c1f4f054c8f893db2a948773164,
title = "Quantitative assessment of local collagen matrix remodeling in 3-D Culture: The role of Rho kinase",
abstract = "The purpose of this study was to quantitatively assess the role of Rho kinase in modulating the pattern and amount of local cell-induced collagen matrix remodeling. Human corneal fibroblasts were plated inside 100-μm thick fibrillar collagen matrices and cultured for 24 h in media with or without the Rho kinase inhibitor Y-27632. Cells were then fixed and stained with phalloidin. Fluorescent (for f-actin) and reflected light (for collagen fibrils) 3-D optical section images were acquired using laser confocal microscopy. Fourier transform analysis was used to assess collagen fibril alignment, and 3-D cell morphology and local collagen density were measured using MetaMorph. Culture in serum-containing media induced significant global matrix contraction, which was inhibited by blocking Rho kinase (p < 0.001). Fibroblasts generally had a bipolar morphology and intracellular stress fibers. Collagen fibrils were compacted and aligned parallel to stress fibers and pseudopodia. When Rho kinase was inhibited, cells had a more cortical f-actin distribution and dendritic morphology. Both local collagen fibril density and alignment were significantly reduced (p < 0.01). Overall, the data suggests that Rho kinase-dependent contractile force generation leads to co-alignment of cells and collagen fibrils along the plane of greatest resistance, and that this process contributes to global matrix contraction.",
keywords = "Actin, Cell mechanics, Collagen matrices, Confocal microscopy, Corneal fibroblasts, Cytoskeleton, Fourier transform, Myosin, Rho kinase",
author = "Areum Kim and Neema Lakshman and Petroll, {W. Matthew}",
year = "2006",
month = "11",
day = "1",
doi = "10.1016/j.yexcr.2006.08.009",
language = "English (US)",
volume = "312",
pages = "3683--3692",
journal = "Experimental Cell Research",
issn = "0014-4827",
publisher = "Academic Press Inc.",
number = "18",

}

TY - JOUR

T1 - Quantitative assessment of local collagen matrix remodeling in 3-D Culture

T2 - The role of Rho kinase

AU - Kim, Areum

AU - Lakshman, Neema

AU - Petroll, W. Matthew

PY - 2006/11/1

Y1 - 2006/11/1

N2 - The purpose of this study was to quantitatively assess the role of Rho kinase in modulating the pattern and amount of local cell-induced collagen matrix remodeling. Human corneal fibroblasts were plated inside 100-μm thick fibrillar collagen matrices and cultured for 24 h in media with or without the Rho kinase inhibitor Y-27632. Cells were then fixed and stained with phalloidin. Fluorescent (for f-actin) and reflected light (for collagen fibrils) 3-D optical section images were acquired using laser confocal microscopy. Fourier transform analysis was used to assess collagen fibril alignment, and 3-D cell morphology and local collagen density were measured using MetaMorph. Culture in serum-containing media induced significant global matrix contraction, which was inhibited by blocking Rho kinase (p < 0.001). Fibroblasts generally had a bipolar morphology and intracellular stress fibers. Collagen fibrils were compacted and aligned parallel to stress fibers and pseudopodia. When Rho kinase was inhibited, cells had a more cortical f-actin distribution and dendritic morphology. Both local collagen fibril density and alignment were significantly reduced (p < 0.01). Overall, the data suggests that Rho kinase-dependent contractile force generation leads to co-alignment of cells and collagen fibrils along the plane of greatest resistance, and that this process contributes to global matrix contraction.

AB - The purpose of this study was to quantitatively assess the role of Rho kinase in modulating the pattern and amount of local cell-induced collagen matrix remodeling. Human corneal fibroblasts were plated inside 100-μm thick fibrillar collagen matrices and cultured for 24 h in media with or without the Rho kinase inhibitor Y-27632. Cells were then fixed and stained with phalloidin. Fluorescent (for f-actin) and reflected light (for collagen fibrils) 3-D optical section images were acquired using laser confocal microscopy. Fourier transform analysis was used to assess collagen fibril alignment, and 3-D cell morphology and local collagen density were measured using MetaMorph. Culture in serum-containing media induced significant global matrix contraction, which was inhibited by blocking Rho kinase (p < 0.001). Fibroblasts generally had a bipolar morphology and intracellular stress fibers. Collagen fibrils were compacted and aligned parallel to stress fibers and pseudopodia. When Rho kinase was inhibited, cells had a more cortical f-actin distribution and dendritic morphology. Both local collagen fibril density and alignment were significantly reduced (p < 0.01). Overall, the data suggests that Rho kinase-dependent contractile force generation leads to co-alignment of cells and collagen fibrils along the plane of greatest resistance, and that this process contributes to global matrix contraction.

KW - Actin

KW - Cell mechanics

KW - Collagen matrices

KW - Confocal microscopy

KW - Corneal fibroblasts

KW - Cytoskeleton

KW - Fourier transform

KW - Myosin

KW - Rho kinase

UR - http://www.scopus.com/inward/record.url?scp=33749157809&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33749157809&partnerID=8YFLogxK

U2 - 10.1016/j.yexcr.2006.08.009

DO - 10.1016/j.yexcr.2006.08.009

M3 - Article

C2 - 16978606

AN - SCOPUS:33749157809

VL - 312

SP - 3683

EP - 3692

JO - Experimental Cell Research

JF - Experimental Cell Research

SN - 0014-4827

IS - 18

ER -