| Cells are known to respond to external stimuli such as chemical, physical and mechanical cues from their microenvironments. In this work, we developed the technology to complement the previously reported studies that deal with substrates' mechanical property by developing thin polydimethlysiloxane (PDMS) membrane microdevices using microfabrication technology. Typically, multiples substrates, each with different stiffness, are used to study the differences in cell's response to the different mechanical properties. With the PDMS membrane devices, a variation in stiffness across a single substrate was generated by strategically integrating micropatterns on a freely suspended PDMS membrane. Human epidermal keratinocytes (HEK) were seeded on the devices and cell migration and accumulation trends, laminin-332 secretion, and focal adhesion and hemidesmosome assembliy were monitored. It was observed that cells within the regions with a small gradient of stiffness were very static while their counterparts on regions with a large gradient of stiffness were extremely active and mobile. This suggests that not only the changes in the property of substrate affect the cells, but the rate at which the changes occur, gradient of stiffness, play a critical role in determining the cells' response. Preliminary data also suggest that HEK specifically secreted more laminin on the softer region of the membrane. Focal adhesion complexes are more prominent on the stiffer region while hemidesmosomes complexes are better developed on the softer region. Concurrently, the same cellular matrix organization on compliant PDMS substrates with Young's modulus ranging from ∼1.2 Pa to 744 kPa was studied. Similar to the membrane devices, more laminin protein secretion and more exptensive hemidesmosome assemblies were observed on the softer substrate while focal adhesions were more prominent, size and number, on the stiffer substrates. We speculate that assembly of hemidesmosome on a less stiff substrate by HEK is typical of normal tissue while enhanced assembly of focal adhesion on stiff substrates reflects what occur when cells interact with the matrix of diseased tissue. |
