Bioactive Surfaces Enabled by Plasma Technology
- Fig. 1: Plasma reactor used for plasma polymer deposition and co-sputtering from a metal target. The plane-parallel electrode set-up enables homogeneous plasma conditions. The gas flow is from the top to the bottom (pumping) side, while the samples are placed on the lower electrode.
- Fig. 2: Cell proliferation (mouse skeletal myoblasts)on a biodegradable, electron-spun scaffold. To enhance cell growth the substrate was coated with a nm-thick oxygen-functional plasma polymer.
- Fig. 3: Characteristics of different Ag release kinetics: 1) initial burst release causing local cytotoxic effects, 2) adjusted Ag release for short-term antibacterial effects (Ag is depleted afterwards), 3) steady state release using gradient layers, and 4) repeated Ag ion releases mediated by degradable layers in a multilayer set-up.
Bioactive surfaces intend to cause a controlled effect at biointerfaces. For this purpose, well-defined properties of the material's surface are required. Plasma technology is well-suited to modify surface properties at the nanoscale, mainly by deposition of ultrathin coatings. Furthermore, a high level of understanding of biological processes at surfaces is essential. Some potential applications of bioactive surfaces are discussed for tissue engineering and antimicrobial effects. Plasma polymer films enable the functionalization of surfaces and immobilization of bioactive compounds as well as the controlled diffusion of drugs.
The aim of bioactive surfaces is to induce a certain, intended response when exposed to bioorganisms. Examples are the support of cell growth, the inhibition of antimicrobial growth as well as the goal to suppress any attachment of bioorganisms, i.e. non-fouling surfaces. While there is increasing success in the first two fields, which are thus discussed in more detail, the latter goal remains challenging, since merely a delay in protein adsorption can be achieved up to date using, e.g., hydrogel-like coatings which hide surfaces by adsorption of abundant water molecules.
Nevertheless, the covalent immobilization of bioactive compounds onto material surfaces (often polymers) enabled considerable progress over past decades in diverse industries as biomedical, bioprocessing, microelectronics, food packaging, and textiles. Many novel life science applications are related to these developments from which, however, also challenges for the materials design occur requiring accurate processing, since precisely tailored surfaces and biointerfaces are mandatory. Plasma technology is thus investigated to functionalize surfaces for the attachment of bioactive compounds and living cells as well as to control the drug release by diffusion barrier layers. Therefore, mainly the deposition of thin films is considered, since film properties can be adjusted over a broad parameter range.
The plasma (as used in materials science) consists of a reactive gas containing ions, electrons, reactive species, and radiation.
By means of electric or electromagnetic fields at high frequencies the plasma is generated at low temperatures (so-called ‘cold‘ plasma) enabling the modification of temperature-sensitive surfaces (Fig. 1). Only a small part of the plasma is made of highly energetic particles causing ablation (etching or sputtering) or densification processes. Thin films can be deposited by the activation of monomers in the gas phase yielding plasma polymerization as well as by sputtering, i.e. the removal of atoms from a target material. Sputtering is mainly used to deposit metal (or metal oxide) films, since high sputtering yields can be achieved at low pressure. For plasma polymerization, different starting monomers (such as hydrocarbons and siloxanes) are used which can also be mixed with inert and reactive gases. Depending on the densification (cross-linking) during film growth highly functional surfaces or dense, hard coatings can be obtained. Plasma polymerization can both be performed at low pressure and atmospheric pressure.
Cell Growth at Surfaces
Cells or cell populations react to a broad spectrum of chemo-, mechano- physico-, and topological signals at a bio-material interface. As scaffold material for cell growth, therefore, a substrate is preferred that mimics the properties of tissue (i.e. tissue engineering). Electrospun, fibrous polymer substrates are of particular interest due to their porous structure, use of biocompatible and/or biodegradable material, and low elastic modulus (comparable to tissue). Polymer surfaces, however, require a further functionalization step in order to obtain a bioactive surface. A simple plasma activation step (e.g. using an oxygen-containing plasma) might thus be used to obtain surface polar groups which, however, readily undergo reorientation processes. Deposition of thin films can avoid such aging effects when they are partly cross-linked (i.e. stabilized) yet still comprising a sufficient functional group density. For the coating of soft, tissue-like substrates the film thickness is limited to a few nanometers, since it has been shown that even plasma coatings exceeding ~10 nm substantially affect the mechanical properties (stiffness) of the substrate due to higher cross-linking. Therefore, ultrathin films are required that, in addition, do not show leaching of oligomeric compounds. Both, oxygen- and nitrogen-functional plasma polymer films have been extensively examined which contain hydroxyl, carbonyl, carboxyl and/or amine groups. While both coatings support cell growth at surfaces, oxygen-functional plasma polymers were found to have a higher stability and better penetration into a 3D-structured substrate. Cells can thus grow and proliferate at such modified surfaces (Fig. 2), which is increasingly used in tissue engineering.
Furthermore, both types of plasma polymers can also be used to covalently bind bioactive compounds (such as bio-linkers, growth factors etc.) at a surface.
To obtain antimicrobial surfaces, mainly two approaches can be distinguished. While the immobilization of antibacterial molecules (such as quaternary ammonium compounds, polyphenols etc.) results in the inhibition of bacteria growth directly at the surface, the release of antibacterial agents also shows an effect on the surrounding media. The first approach is interesting for medical devices, displays, food packaging, and textiles. Again functional plasma polymer layers might be used for the immobilization of the bioactive compounds. The second approach based on drug release is mainly considered for wound care, catheters, sanitation, and implants. Here, mainly silver is used as the antibacterial agent due to its efficacy against a broad spectrum of bacteria and fungi. Furthermore, compared to copper or zinc oxide, silver (Ag) shows the lowest risk for human beings and the environment. The antibacterial effect is related to the release of Ag ions in aqueous media which interact with the metabolism, the respiration and replication system of microorganism. Nevertheless, high concentrations of Ag ions can also cause cytotoxic effects, i.e. cell populations become affected. Ag-rich surfaces typically yield an initial burst release of Ag ions which results in locally cytotoxic conditions (Fig. 3). To avoid such conditions but still enable antibacterial efficacy, the Ag ion release has to be adjusted. Using plasma polymerization, diffusion barriers can be deposited either on Ag-rich surfaces or during co-sputtering from a silver target. Gradients in the Ag content, i.e. more Ag in depth and less towards the surface, enable a steady Ag ion release. Some applications, e.g. implant surfaces, require a short-term Ag ion release or a recurrence of released silver over longer timeframes (see Fig. 3). Adjusted, Ag-poor coatings or multilayers of Ag-containing layers with degradable plasma polymers can fulfill the latter purpose.
While there is substantial progress in the field of antibacterial surfaces by fine tuning of the material's properties, different strains of the same bacteria were found to give a different response for the same antibacterial test conditions. Antibacterial tests should thus be repeated with different strains in order to enhance the reliability of the intended efficacy of a bioactive surface. Moreover, further understanding of the involved biological processes is required.
Plasma technology has been shown to support cell growth, to immobilize bioactive compounds as well as to control the efficacy of antibacterial surfaces by avoiding cytotoxic effects. Beside the discussed contributions in tissue engineering and for antibacterial surfaces, bioactive surfaces are of growing interest for diverse fields such as biochips, biosensors, drug delivery, bioseparation, cell engineering, and stem-cell differentiation.
In order to obtain reliable materials and processes yielding well-defined bioactive surfaces, specialists from both fields of materials science and biology should further strengthen their collaborations. On one side, the performance of modified surfaces in a biological environment is under investigation, where plasma technology becomes an increasingly important tool, while on the other side, biologists steadily improve their know-how about important processes at biointerfaces. Further progress in life science applications can thus be expected.