Elsevier

Acta Biomaterialia

Volume 6, Issue 10, October 2010, Pages 3824-3846
Acta Biomaterialia

Review
The interaction of cells and bacteria with surfaces structured at the nanometre scale

https://doi.org/10.1016/j.actbio.2010.04.001Get rights and content

Abstract

The current development of nanobiotechnologies requires a better understanding of cell–surface interactions on the nanometre scale. Recently, advances in nanoscale patterning and detection have allowed the fabrication of appropriate substrates and the study of cell–substrate interactions. In this review we discuss the methods currently available for nanoscale patterning and their merits, as well as techniques for controlling the surface chemistry of materials at the nanoscale without changing the nanotopography and the possibility of truly characterizing the surface chemistry at the nanoscale. We then discuss the current knowledge of how a cell can interact with a substrate at the nanoscale and the effect of size, morphology, organization and separation of nanofeatures on cell response. Moreover, cell–substrate interactions are mediated by the presence of proteins adsorbed from biological fluids on the substrate. Many questions remain on the effect of nanotopography on protein adsorption. We review papers related to this point. As all these parameters have an influence on cell response, it is important to develop specific studies to point out their relative influence, as well as the biological mechanisms underlying cell responses to nanotopography. This will be the basis for future research in this field. An important topic in tissue engineering is the effect of nanoscale topography on bacteria, since cells have to compete with bacteria in many environments. The limited current knowledge of this topic is also discussed in the light of using topography to encourage cell adhesion while limiting bacterial adhesion. We also discuss current and prospective applications of cell–surface interactions on the nanoscale. Finally, based on questions raised previously that remain to be solved in the field, we propose future directions of research in materials science to help elucidate the relative influence of the physical and chemical aspects of nanotopography on bacteria and cell response with the aim of contributing to the development of nanobiotechnologies.

Introduction

While the application of nanotechnology in the fields of electronic and optical engineering is well established, the use of nanostructured materials in other fields, such as medicine and biology, is truly novel. The field of biological micro-electromechanical systems (bioMEMS) and their associated biomedical nanotechnologies are developing at a rapid pace. The most notable applications are biomolecular and cell analysis, microfluidics, biomedical drug delivery and medical prostheses. In several of these fields an understanding of cell–substratum interactions is the key to the future development of these technologies.

Tissue engineering is generating increasing interest as results obtained in cells associated with engineered materials have shown promise for the restoration of function to damaged tissues. Again, a crucial point in this field is understanding cell–material interactions. The scaffolds can be considered at different length scales: the macrostructure, the microstructure and the nanostructure. On the macro and micro scales it has been shown that surface chemical composition and surface topography have strong effects on cell behaviour [1], but less is known about how cells react to nanoscale structures. However, cells are likely to be able to respond to nanostructures, since in vivo they live inside an extracellular matrix (ECM) containing nanoscale collagen fibrils and since their own surface is structured on the nanoscale level (receptors and filopodia). In the last few years methods for the fabrication of nanoscale controlled topographies have been developed and can now be used to tailor surface chemistry and surface topography in order to elucidate how cells respond to nanotopography. However, the major problems researchers interested in cell/material research at the nanoscale level face are the limits of microscopical resolution and those associated with producing large areas of nano-fabricated materials (cost and time). Initial studies have shown that cells are able to respond to nanostructures, but more work is needed to comprehend the mechanisms underlying cell responses, in particular how to design a surface to induce a desired behaviour, which is essential to improve the design of medical implants and for tissue engineering approaches. The first essential step in such a study is the controlled fabrication of model surfaces. Thus, this review will start with a description of the techniques available for physical and chemical patterning. The influence of nanotopography on surface chemistry and methods available to analyse surface chemistry at the nanoscale will be discussed. The main focus of this review will be to describe current knowledge on eukaryotic cell responses to surfaces presenting a controlled nanotopography. We have chosen to focus on studies in which at least one dimension is on the nanometre scale: lateral or vertical for surface topography or lateral resolution for chemical topography. The influence of the size and shape of nanofeatures, as well as the spacing and the organization of nanofeatures on cell response will be presented. Since cells have to compete with bacteria for the surface in many environments, and the prospect of using topography to create bacteria-repellent surfaces is important for tissue engineering, the limited current knowledge on this topic will also be reviewed in this paper. The role of protein adsorption on nanotopography and its resulting effect on cell or bacteria will also be described. Finally, we will review current and upcoming technologies that use cell–surface interactions at the nanoscale and propose future directions for research in this domain.

Section snippets

Patterning at the nanometre scale

Surface patterning is a field in which the resolution limit is constantly being improved. The widespread development of microfabrication techniques and the development of self-assembly techniques are such that the production of structures on the sub-micron scale is now becoming routine. Two types of surface patterning should be distinguished: structural topography and chemical patterning. At the nanometre scale it is sometimes difficult to separate the two and often techniques which are used to

How does nanotopography influence surface energy?

One important point to remember is that although the nanotopographied surface created is chemically similar to the bulk, the nanotopography itself may induce surface chemistry or energy differences.

One well-known example is the increase in hydrophobicity of a microstructured surface known as the lotus effect. The presence of microstructures effectively increases the air content of the surface, which increases the surface contact angle, as described by the Cassie–Baxter equation [52], [53]. This

How can surface chemistry be characterized at the nanometre scale?

The physical characterization of surface chemistry at nanometre scale lateral resolution has reached some maturity, with several techniques now widely available. For an analysis method to be applicable in the assessment of functional nanostructures and nanopatterns on solid surfaces the technique needs to fulfil some essential conditions: it must be highly surface-sensitive (to be able to collect a signal from a very small volume) and surface-specific (the signal should correspond only to the

How does a cell interact with a surface at the nanometre scale?

Globally, the way cells interact with a surface presenting nanostructures is the same as at other scales. However, the interface of a cell with a surface consists of discrete attachment points [1]. Several biological molecules can act as cell attachment molecules, but the most common are the integrins (Fig. 1). Integrins are heterodimeric proteins which are constituted of two subunits, α and β. Upon attachment to a surface the integrin receptors cluster together and recruit cytoplasmic proteins

How do bacteria interact with surfaces?

Bacteria are prokaryotic cells and differ from eukaryotic cells in several aspects. Their cell wall is composed of phospholipids, like eukaryotic cells, but they are much more rigid. This is in part due to an external layer of peptidoglycan, which is thick in Gram-positive bacteria whereas Gram-negative bacteria have a thin peptidoglycan layer which is covered by an additional polysaccharide outer layer. They vary greatly in size (from under 1 μm to several tens of microns) and shape (spherical,

Protein adsorption on nanotopography: its role in cell and bacterial responses to nanotopography

It is well established that implanted materials are immediately coated with proteins from the blood and interstitial fluid, and it is through this adsorbed layer that cells sense foreign surfaces. It is the adsorbed proteins, rather than the surface itself, to which cells initially respond [204]. Thus, it is of fundamental interest to understand how the properties of nanostructured materials influence the structure, activity and stability of conjugated proteins. Additionally, the retention of

Applications

As we have previously seen, the influence of nanotopography alone, or combined with nanoscale chemistry, has been under intensive investigation. Although the mechanisms by which the substrate topography modifies cell behaviour – in areas ranging from adhesion, growth and motility to differentiation – are beginning to be understood, their integration in finished devices is still in the development phase.

One exciting prospect is given by results obtained for the differentiation of bone cells. A

Future directions

Several research pathways emerge from this review of the most recent knowledge of how cells respond to nanostructured materials. First of all, a major point in cell responses to surfaces appears to be the role of proteins adsorbed on the surfaces. Proteins, because of their size, are particularly sensitive to nanotopography and to chemical topology at the nanometric scale. The future of research in this field will certainly rely on a better analysis and understanding of the influence of

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