The mechanics of nanomedicine

This essay was written by Tania Saxl and was first published in the 2011/12 Mill Hill Essays.

If you attempt to look inside the tissues of the body and examine the fundamentals of disease, you must enter the nano-world. This is the 1-1000 nanometre range, which is bigger than a few atoms but smaller than a human blood cell, and is the scale at which much of human biology takes place. The nanoscale is therefore of huge importance to medicine. Viruses such as HIV and influenza are invisibly small nanoparticles that attack cells and replicate using nano-mechanics. Nanoscale electrical networks enable the brain to process information and the heart to maintain its beat. DNA is wound up in nano-structures, and read by nano-machines. Alzheimer’s disease and Parkinson’s disease are caused by nanoscale fibres. Tuberculosis and pneumonia are caused by bacteria, which use complex nano-motors such as tail-like flagellae and pili to invade the body.

One area of nanomedicine covers the use of nano-sized drugs for therapy and imaging, but it also concerns ways to exploit mechanical and electronic devices on the nanoscale. Using nanotechnology to develop tools to visualize biological processes and to engineer medical devices is especially valuable for the future of medicine. This essay will look at some of the technologies being developed in nanomedicine and their potential uses.

The nano-toolbox

The nanoscale is difficult to comprehend, as you cannot see things nano-sized by eye, or even using a conventional microscope. Nanoscale objects, such as a virus, are just too small. The following section describes a few of the important techniques that can be used to image and manipulate on this scale.

Atomic Force Microscopy (AFM)

Tools for achieving resolution at the scale of atoms were originally designed for the semiconductor industry but are now being applied to the molecules of life. The ScanningTunneling Microscope (STM) was developed by Gerd Binnig and Heinrich Rohrer, scientists at IBM in Switzerland, and they won the Nobel Prize in 1986 for their work. The STM was famously used to write the letters ‘IBM’ in individual silicon atoms, a breakthrough in the ability to control the building blocks of matter. Later the Atomic Force Microscope (AFM), a derivative of the STM, was developed and is now used in biological research. It works by vibrating a tiny silicon lever with a sharp silicon tip, close to the surface under investigation. Changes in this vibration caused by surface topography are detected using a laser. The smaller the tip and the faster the vibration, the smaller the features that can be observed.

The nano-toolbox
Figure 1. The nano-toolbox

It is now possible to image the molecular structure of DNA and its surface charge with nanometre resolution, which helps us understand how all our DNA packs into each cell nucleus. The AFM can also be used to image nano-sized pores on the outside of our cells. These pores, called membrane channels, selectively allow ions or small molecules in and out of the cell, and some hereditary diseases such as frontal lobe epilepsy and cystic fibrosis are associated with their malfunction. They are particularly difficult to study, as they tend to fall apart if you try to remove them from the membrane. Whereas other techniques often require drying or crystallising, a high frequency AFM is able to image surfaces in liquid so that the pores and their mechanism can be explored in a natural state.

Focused Ion Beam Scanning Electron Microscopy (FIBSEM)

FIBSEM is a combination of two techniques. A beam of charged atoms is used to physically slice away nano-thin layers of a specimen as they are imaged using an electron microscope. The structure is peeled away one layer at a time, to reveal a three dimensional image. This is ideal for imaging the network of connections between the cells that make up the brain, the most complex organ of the body. The cells of the brain are connected in a dense network, and the research discipline known as connectomics aims to map the detailed wiring of the brain’s circuits. It is extremely difficult to follow the pathways of nerve cells in the brain because they are so small and thin. Reconstructing them in three dimensions is a delicate task and images take weeks to acquire. With FIBSEM, it is possible to image neural circuits at a resolution of a few nanometres in all three dimensions. Disentangling the network of neural cells and their connections will enable the link between structure and function in the brain to be explored, which is vital in the quest to understand neurological disorders.

3D Neural circuit imaging using FIBSEM
Figure 2. 3D Neural circuit imaging using FIBSEM.

Resolution approximately nine nanometres. In the right-hand image, individual cells are identified by outlining in yellow.

Image courtesy of Arnd Roth and Michael Hausser, (London Centre for Nanotechnology and Wolfson Institute for Biomedical Research, University College London)

Coherent X-ray diffraction

X-ray crystallography has been the workhorse of biological imaging since the time when Rosalind Franklin produced diffraction patterns corresponding to the double helix of DNA. It relies on being able to produce a crystal from the molecule to be imaged, an often tricky and lengthy exercise. A modern adaptation of this technique, the imaging of single molecules or complexes of molecules, is coherent X-ray diffraction. The ability to image single complexes with nanometre resolution requires the use of a synchrotron facility. This is a machine, several football pitches in size, that can accelerate electrons to near the speed of light. From the movements of these electrons a bright beam of X-rays is released, sufficiently bright and ‘coherent’ or ordered, to image the molecule under investigation. Already used to image the interactions between biological molecules and metal surfaces, fundamental to the fabrication of biosensors, these bright and uniform X-ray beams could be used to show the make-up of our own chromosomes. This could help us to understand genetic diseases that are caused by chromosome defects, and to study the mechanism by which the chromosome is formed and read.

Electron Paramagnetic Resonance Spectroscopy (EPR)

EPR is a type of spectroscopy that detects free radicals, that is, molecules with ‘free electrons’. The state of these free radicals can provide information on their surrounding environment. Some biological molecules that contain free radicals are fundamentally important, such as the proteins, small molecules and vitamins involved in respiration and metabolism. EPR allows a researcher to observe the processes involved in respiration, for example, by tracking the free radicals. This is especially powerful when combined with static information, such as crystallography, to give a picture of the dynamic process.

Some things that are not naturally detectable by EPR can be made so by attaching a ‘spin label’ to them, that is, a small molecule with a free radical part.This was used to study the mechanism by which harmful strains of Escherichia coli infect a human. E. coli invades by attaching to the lining of your intestine. If the bacteria cannot stick, they are rendered harmless.The bacterium uses thin hair-like filaments, called pili, to form these attachments to the intestine wall. Although it is possible to see an E. coli cell under a high-power microscope, the pili are only a few nanometers wide, and are beyond the resolution limit of a light microscope. New pili are constantly grown by the bacteria, which they assemble one unit at a time. Understanding how formation of the pili occurs can help us to prevent infections. Crystallography allows us to examine the pilus in a ‘frozen’ crystallised state. When EPR is used in combination with crystallographic images, the formation of the pilus, one step at a time, can be pictured.

Doppler interferometry

Many processes in biology involve not only tiny nano-structures but also nanoscale motion of these structures, which can be extremely rapid. For example, the motion of sound-detecting ‘hair bundles’ in the ear requires a special technique to detect their tiny and rapid fluctuations. Sound waves cause hair bundles in the ear to vibrate.These hair bundles are microscopically small and vibrate with invisible nanoscale deflections, which occur hundreds or thousands of times a second.This delicate movement can be detected using a laser beam focused on the tip of the hair bundle, and the pattern of reflected laser light is used to measure the rapid movement of the structure.

Understanding the mechanical processes of hearing and how the sound gets transduced into an electrical signal can help with the design of cochlear implants, synthetic hearing devices which artificially stimulate the nerves.

Nanotechnology for diagnostics in healthcare. Gold nanoparticles and films

Most new types of diagnostic sensor for healthcare are made from thin gold layers or small gold spheres, this is due to the ease with which you can attach biological molecules to gold. The smaller these devices are made, the more sensitive they become, therefore it is important to examine on the smallest scale. Gold nano-particles have been studied using the nano-scopic technique of coherent X-ray diffraction. It was found that an individual nanometre-sized grain of gold becomes distorted when a single layer of molecules is attached, such as those that would be used in sensing. As sensors get smaller to increase sensitivity, effects such as these become more significant and understanding nanoscale behaviour becomes increasingly important when developing new diagnostics.

Surface Plasmon Resonance

Optical techniques such as Surface Plasmon Resonance (SPR) can be used as a sensor to analyse blood and other liquids. Nano-sized metal features, such as gold drops on glass, are irradiated with laser light. This causes a thin, invisible electromagnetic field to be created on the gold surface.This delicate field is easily disrupted by the arrival of a protein or an interaction between a protein bound to the surface and a drug, and can therefore be used to diagnose infection or to screen for novel drugs.

Cantilevers

One approach to detecting nanosized species and studying nanoscale interactions is the bending of ‘cantilevers’. Silicon levers the width of a human hair are arranged like a series of diving boards in a row. These levers are coated with receptor molecules, which can bind to bacterial cells, viruses and proteins. Nanometre bending of these tiny levers is detected using a laser.

Infectious bacteria such as Methicillin-resistant Staphylococcus aureus (MRSA) are becoming increasingly problematic in healthcare.They are resistant to most existing antibiotics and therefore new drugs are needed to combat them. Critical to finding the correct drug is an understanding of the mechanism by which it works. To investigate new antibiotics for MRSA, cantilevers have been coated with MRSA specific proteins. An antibiotic interaction with the proteins bends the levers, due to stress. Experiments such as these can tell us which drug binds most effectively to MRSA, indicating its potential to treat the disease.

The cantilevers are not limited to detecting drugs; the size range is relatively wide, from whole organisms, such as the tuberculosis bacterium, down to tiny HIV virus particles and HIV-specific proteins. Through using cantilevers, it is possible to detect disease and to understand better the mechanism of infection and potential cure.

Mechanical cantilever sensors for detecting markers of disease.
Fig 3. Mechanical cantilever sensors for detecting markers of disease.
Image courtesy of the London Centre for Nanotechnology

Particle sorting

Often when diagnosing a viral infection or reacting to an epidemic outbreak it is necessary to isolate the virus and find out what it contains as fast as possible. Virus particles themselves are only few nanometers in size, invisible under a microscope and often in such small quantities in the blood that only a few hundred are hidden amongst the billions of much larger cells in a sample. In order to detect these, the viruses have to be found, sorted and concentrated, a task for a nano-engineered device. A forest of nano-pillars can be used as an obstacle course that causes particles flowing through it to be deflected according to size. Thus the hidden viruses can be separated from the sea of red and white blood cells and collected. This type of sorting is quite remarkable, and can also be used to make viral vaccines, where the tiny viruses must be delicately sifted from the host in which they are grown.

Engineering future implantable medical devices

Many medical procedures involve the implantation of metal or other non-biological structures and devices within the body. The engineering and design of these implants can be critical to their performance, the nanoscale properties in particular can have a large impact on interaction with cells and molecules in the body.

Designing surface properties

Nanotechnology often takes its inspiration from nature. Researchers developing a super-hydrophobic surface, one that is extremely repellent to water, have copied the nanoscopic structure of the lotus leaf. The surface of a lotus leaf is rough and waxy, covered in tiny wax pyramids that are invisible to the eye. By mimicking this design, it is possible to engineer a coating that causes water to form perfectly spherical droplets on its surface, which then roll off at the slightest movement. This coating has possible application as an antithrombotic coating for medical implants, to allow the streamlined passage of blood and discourage sticking and clogging.

Nanoscale electrode arrays

Receiving and transmitting electrical signals is fundamental to many processes in the body. Our heart beats to an electrical rhythm, our brain operates electrically, our muscles twitch using electrical impulses and we are able to see because images are converted to and transmitted as a series of electrical pulses. In order to measure and understand these signals, and to replace and repair damaged function, we can use electrical devices. The smaller and more biocompatible the device, the better it can integrate with the body. This is where nanotechnology plays a role.

Stimulation of Ganglion cells in the retina of the eye causes the brain to form images of the outside world. Micro-electrode arrays are being developed to be implanted in the eye, replacing damaged cells and stimulating the eye to form an image. Creating an intimate contact between the cells in the eye and the electrodes is a delicate task, as these cells in particular don’t like to grow on electrodes. By understanding and manipulating the nanostructure of the electrodes, cells can be encouraged to make contact with the electrodes and form a natural contact with the foreign device. An engineered material, such as nanocrystalline diamond, offers both the desired electrical properties, as it can transmit tiny electrical signals to the cells, and is also made up of features that encourage the cells to grow on its surface.

Engineering cell growth

Novel materials can be designed to support and help regrowth of cells. Some cells are very fussy about surfaces, and will only grow and thrive when the surface has a specific nano-topography. By controlling the nanoscopic shape and chemical properties of a cell’s environment, patterns of growth can be stimulated. This technique has been used with success on the surface of hip implants. Neurons, the cells responsible for the transmission of information in the brain, are notoriously difficult to re-grow after damage. Initial experiments indicate that a carefully engineered surface such as nano-crystalline diamond can be as effective as a protein treated surface designed specifically for neurons.

Application: improved cardiac stents

A good example of a medical device that can benefit from nano-engineering is the stent. A stent is a small tube of steel mesh, commonly used to open up arteries in the case of blockage. Such blockages are often caused by the build-up of fatty layers and plaques inside the arteries of the heart. The stent can be expanded inside the artery, concertina-like, to increase the size of the opening and to hold it open. The surface of the stent or the stent material itself can be nano-engineered to provide useful properties. Implantation of a stent carries a risk of clotting and the cells of the artery are usually damaged leaving the wall weak. In the future, superhydrophobic coatings (mentioned previously) may reduce clogging and thrombosis. Surface patterning or coating with a nano-engineered material such as nanodiamond may encourage the weakened cells of the epithelium to regrow at the region of the implant.

Conclusion

Physics and engineering techniques can be applied to the medical field in order to allow us to interact with the nanoscale. Tools developed for the semi-conductor industry and nano-materials research, to study from atoms to complex systems, can be applied to open up the biological world to the surgeon, doctor and medical scientist. These tools complement traditional medical methods and enable new possibilities in diagnosis and therapy.

The London Centre for Nanotechnology (LCN) is a joint venture between UCL and Imperial College London.A multidisciplinary centre at the forefront of science and technology, the purpose of the LCN is to solve global problems in healthcare, IT and the environment through the application of nanoscience and nanotechnology. Experimental research is supported by world leading modelling, visualisation and theory, and through access to state-of-the-art cleanrooms, design, fabrication and characterisation laboratories.The LCN is a partner to industry.
www.london-nano. com

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