W. Richard Bowen and Nidal Hilal 4
W. Richard Bowen and Nidal Hilal 4
W. Richard Bowen and Nidal Hilal 4
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7.2 ENgINEERINg THE ECM FOR PRObINg CELL SENSINg 203<br />
of parenchymal cells such as hepatocytes (liver cells) which require heterotypic<br />
interactions between non-parenchymal cells (e.g. fibroblast) to<br />
maintain their liver cell phenotype [30]. In this study, the microfabrication<br />
approach allowed researchers to specify independent variables, including<br />
the formation of a heterotypic interface <strong>and</strong> the ratio of cell populations<br />
at specific locations in their samples, something which was not possible<br />
using traditional r<strong>and</strong>om co-culture methods.<br />
Although many of the above findings have been made possible with<br />
the aid of micropatterning, they have also indicated the desirability of<br />
further investigation at smaller length scales (�100 nm) where most of the<br />
molecular mechanisms relevant to cell biology can be discovered. Thus,<br />
we describe relatively recent investigations that have used nanopatterned<br />
engineered surfaces in the next subsection.<br />
Nanopatterning<br />
To generate nanopatterns, electron beam lithography (EBL) is normally<br />
used since the spatial resolution of photolithography is limited<br />
by the diffraction of light. Rather than using a mask, EBL uses a focused<br />
electron beam to directly write patterns onto an electron beam sensitive<br />
resist. Since this is a serial writing method, i.e. tracks are written segment<br />
by segment, this technique can require a significant amount of time to<br />
write a single large area pattern <strong>and</strong> is thus very expensive. However, in a<br />
development similar to the soft lithography described earlier, nanopattern<br />
structures can also be imprinted onto a solid polymeric substrate (nanoimprinting<br />
lithography [NIL]), greatly reducing the cost [31, 32]; Figure 7.4).<br />
By combining NIL with self-assembled monolayer techniques, protein<br />
nanopatterns of dimension �100 nm have been produced [33, 34]. The<br />
combined capability of NIL <strong>and</strong> EBL for the generation of arbitrary nanopatterns<br />
on a wide range of materials has also led to the discovery of cell<br />
response to nanotopography, as discussed in later sections.<br />
Other methods for nanopatterning biological molecules include scanning<br />
probe lithography [35], self-assembly nanofabrication using block<br />
copolymer, <strong>and</strong> colloidal lithography [36]. A good review of these techniques<br />
is given by Gates et al. [37]. However, to generate a statistically<br />
meaningful cellular study, fine tailored adhesive nanopatterns have to<br />
cover a large area, preferably of the order of square centimetre. This<br />
imposes a big challenge for some of the serial writing methods, such as<br />
scan probe-associated lithography, although new developments in parallel<br />
writing using multiple tips might mitigate this barrier.<br />
As an example of an extension of the self-assembled block copolymer<br />
technique, recently, Spatz’s group have developed the ‘micelle diblock<br />
copolymer lithography’. This allows precise control of space between<br />
RGD lig<strong>and</strong>s at the length scale of 10–200 nm [38]. This strategy uses selfassembly<br />
of diblock polymer of polystyrene-block-poly(2-vinylpyridine)