Submitted on September 7, 2005
Accepted on January 8, 2006
The Intrinsic Mechanical Properties of the Extracellular Matrix Affect the Behavior of Pre-Osteoblastic MC3T3-E1 Cells
Chirag B Khatiwala1, Shelly R Peyton1, and Andrew J Putnam2*
1 Chemical Engineering and Materials Science, University of California, Irvine, Irvine, CA, USA
2 Chemical Engineering and Materials Science, University of California, Irvine, Irvine, CA, USA; Biomedical Engineering, University of California, Irvine, Irvine, CA, USA
* To whom correspondence should be addressed. E-mail: aputnam{at}uci.edu.
Mechanical cues present in the extracellular matrix have been hypothesized to provide instructive signals that dictate cell behavior. We probed this hypothesis in the context of osteoblastic cells by culturing MC3T3-E1 cells on the surface of type I collagen-modified hydrogels with tunable mechanical properties and assessed their proliferation, migration, and differentiation. On gels functionalized with a low type-I collagen density, MC3T3-E1 cells cultured on polystyrene proliferated twice as fast as those cultured on the softest substrate. Quantitative time-lapse video microscopy analysis revealed random motility speeds were significantly retarded on the softest substrate (0.25 ± 0.01 µm/min), in contrast to maximum speeds on polystyrene substrates (0.42 ± 0.04 µm/min). On gels functionalized with a high collagen-I density, migration speed exhibited a biphasic dependence on ECM compliance, with maximum speeds (0.34 ± 0.02 µm/min) observed on gels of intermediate stiffness while minimum speeds (0.24 ± 0.03 µm/min) occurred on both the softest and most rigid (i.e., polystyrene) substrates. Immature focal contacts and a poorly organized actin cytoskeleton were observed in cells cultured on the softest substrates, while those on more rigid substrates assembled mature focal adhesions and robust actin stress fibers. In parallel, focal adhesion kinase activity (assessed by detecting pY397-FAK) was influenced by compliance, with maximal activity occurring in cells cultured on polystyrene. Finally, mineral deposition by the MC3T3-E1 cells was also affected by ECM compliance, leading to the conclusion that altering ECM mechanical properties may influence a variety of MC3T3-E1 cell functions, and perhaps ultimately, their differentiated phenotype.
