Page 41 - Nanotechnology an introduction

grooves. The reader appraising this literature should be warned that sometimes these surfaces are referred to as “nano” without any real
justification (since the features are larger than nanoscale).
The goal of investigations of the nano/bio interface could be to demonstrate that unique features of cell response arise as feature sizes fall below
100 nm (taking the consensual definition of the upper limit of the nanoscale). Surprisingly, this has hitherto been done in only a very small number of
cases. One of the few examples is the work by Teixeira et al. [160], who showed that when the ridges of a grooved pattern were shrunk to 70 nm,
keratocyte alignment was significantly poorer than on ridges 10 times wider (whereas epithelial cell alignment was unchanged). Alternatively,
following the spirit of Chapter 2, one could define one or more “biological response nanoscales” by presenting cells with a systematically
diminishing set of structural features and observing at what scale the cells started to respond. Alignment is of course only one, perhaps the most
trivial, of the huge variety of measurable responses. Table 4.1 summarizes some of the others.

A rather diverse set of observations may be systematized by noting that the fundamental response of a cell to an environmental stimulus is an
adaptive one. Following Sommerhoff [154], one can distinguish three temporal domains of adaptation: behavioral (short-term); ontogenic (medium-
term); and phylogenetic (long-term). Although Sommerhoff was discussing organisms, these three domains have their cellular counterparts. At the
cellular level, behavioral adaptation is largely represented by energetically driven phenomena [101], perhaps most notably the spreading transition
characteristic of many cells adhering to substrata (Figure 4.3). Morphological changes have been shown [91] and the rate of spreading has been
quantitatively shown, to depend on nanoscale features of the substratum [8].

Figure 4.3 Idealized cell (initial radius R) spreading on a substratum F in the presence of medium C. Upper panel: cross-sections, showing the transition from sphere to segment (h is the ultimate height of the cell). Lower panel: plans, as
might for example be seen in a microscope (the dark region shows the actual contact area).
The secretion of microexudates, growth factors, etc. would appear to involve the reception of chemical and/or morphological signals by the cell
surface and their transmission to the nucleus in order to express hitherto silent genes. Indeed, this secretion appears to be part of an adaptive
response to condition an uncongenial surface. Work in molecular biology has established that the amino acid triplet arginine–glycine–aspartic acid
(RGD), characteristically present in extracellular matrix proteins such as fibronectin, is a ligand for integrins (large transmembrane receptor
molecules present in the cell membrane), binding to which triggers changes depending on cell type and circumstances. Such phenotypic changes
fall into the category of ontogenic adaptation, also represented by stem cell differentiation. Recently reported work has shown that 30-nm diameter
titania nanotubes promoted human mesenchymal stem cell adhesion without noticeable differentiation, whereas 70–100-nm diameter tubes
caused a ten-fold cell elongation, which in turn induced cytoskeletal stress and resulted in differentiation into osteoblast-like cells [126].
Phylogenetic adaptation involves a change of genetic constitution, and would correspond to, for example, the transformation of a normal cell to a
cancerous one. There are well-established examples of such transformations induced by certain nanoparticles and zeolites. Two important criteria
for deciding whether a nanoparticle might be carcinogenic seem to be:

1. Whether the particle is insoluble (this does not, of course, apply to nanoparticles made from known soluble carcinogens active when dispersed
molecularly).
2. Whether the particle is acicular and significantly longer than macrophages (see Figure 4.3).
If both these criteria are fulfilled the macrophages ceaselessly try to engulf the particles and they become permanent sites of inflation, which may be
the indirect cause of tumorigenesis. Nevertheless, given the long induction periods (typically decades) that elapse between exposure to the
particles and the development of a tumor, elucidation of the exact mechanism remains a challenge. Incidentally, there appear to be no known
examples of nanostructured substrata that directly induce cancer through contact. However, it should be kept in mind that the mechanism for any
influence may be indirect (as in [126]; note also that an extracellular matrix protein might bind to a certain nanotextured substratum, change its
conformation—see Section 4.1.4—and expose RGD such that it becomes accessible to a cell approaching the substratum, triggering ontogenic
changes that would not have been triggered by the protein-free substratum) and is generally unknown. The secretion of macromolecular,
proteinaceous substances that adsorb on the substratum is an especially complicating aspect of the nano/bio interface, not least because any
special nanostructured arrangement initially present is likely to be rapidly eliminated thereby.

Prokaryotic Cells
In contrast to eukaryotes, prokaryotes (archaea and bacteria) are typically spherical or spherocylindrical, smaller (diameters are usually a few
hundred nm) and enveloped by a relatively rigid cell wall predominantly constituted from polysaccharides, which tends to maintain cell shape.
Prokaryotes have received rather less attention than eukaryotes, although the interaction of bacteria with surfaces is also a topic of great
importance for the biocompatibility of medical devices, which is normally viewed in terms of adhesion rather than adaptation. Nevertheless, if one
considers bacterial communities, the formation of a biofilm, based on a complex mixture of exudates, and which usually has extremely deleterious
consequences, should be considered as an adaptive response at least at the ontogenic level, since the expression pattern of the genome changes
significantly. A worthy (and hitherto unreached) goal of investigation, therefore, is whether one can prevent biofilm formation by nanostructuring a
substratum.

4.1.4. Biomolecules
A protein is comparable in size or smaller to the nanoscale features that can nowadays be fabricated artificially. The problem of proteins adsorbing
to nonliving interfaces has been studied for almost 200 years and an immense body of literature has been accumulated, much of it belonging to the