Creating a window into the body

17 Jun 2002

Researchers in the US and Russia have pioneered techniques for optical clearing of tissue that offer many potential advantages in laser surgery and phototherapy.

The idea is that by reducing the scattering effect of the tissue, imaging and surgical techniques can probe deeper into the body than is normally possible. This effectively opens a window into the body for more effective treatment, aiding diagnosis of deeper-lying tumours, for example.

All tissue scatters visible and near-infrared light far more effectively than it absorbs it. This reduces the contrast between normal and diseased tissue, such as tumours, when imaged. There are two ways to tackle this problem: either increase the absorption of the target, or reduce the scattering of the surrounding tissue. The first option is tackled by photosensitizing drugs: these increase absorption, such that light can be more effectively used in treatment. One problem associated with such treatment is the toxicity of the photosensitizers, and a major advantage of the clearing agents is that they are generally much less harmful.

The key to reducing tissue scatter is to index-match the various components that the tissue is constructed from. Normal tissue has a high water content and there is a large discrepancy between the refractive indices of tissue fibre and water. One way to reduce this discrepancy is to replace the water with an agent that has a better refractive-index match. Ashley Welch of the University of Texas recently made news by doing this in a very simple way. He injected glycerol into rat and hamster skin. Glycerol replaces water in the tissue by osmosis, thereby improving the index matching. The result is a clearing of the tissue, or as some have dubbed it transparent skin (see image sequence in figure 1).

The use of chemicals like glycerol to clear tissue is not new. Over a decade ago, Valery Tuchin's group at Saratov State University, Russia, used the same technique to clear rabbit-eye tissue. Since then, it has performed histological studies on a range of other substances, including glucose and propylene glycol and extended the technique to the human eye (figure 2). Different tissues demand different agents for the clearing process to work effectively. Tuchin explained: "Small-molecule agents, such as glucose, can penetrate tissue, and even cells, whereas large, polymeric agents like polyethylene glycol cannot. To clear the sclera (eye tissue), we must use the smaller agents, as the larger ones merely dry out the tissue."

Tissue clearing is a reversible process. After a short time, typically 20 to 30 minutes, water is reabsorbed into the tissue and the scattering returns to normal. The process can also be actively reversed by immersing the sample in saline solution, which disperses the clearing agent.

When tissue is cleared, diagnostic techniques, such as optical coherence tomography, can produce much clearer, deeper images. Laser microsurgery of blood vessels under the skin is improved due to the reduced scattering, which keeps the laser beam sharp and its energy delivery highly localized. For advanced photodynamic therapy, combinations of absorption-increasing and tissue-clearing agents can be formulated.

Tuchin's group, along with Welch's, lead the world in the field of tissue clearing. At the recent SPIE conference on biomedical optics in Amsterdam, there was a large number of papers and posters presented by Tuchin's 20-strong team. Saratov's collaborations include Huazhong University in China (in vivo rat skin), Ricky Wang's Keele group in the UK (blood vessel and internal organ imaging) and Britton Chance's University of Pennsylvania team (skin and eye applications).

Saratov researchers have studied a wide range of tissues, including muscle, brain and cartilage, as well as internal organs, such as the stomach and colon. They have also induced clearing using drops of the agent. "The challenge now is to find out for how long we can apply the agent without destroying cells or the fibre structure," Tuchin told Opto and Laser Europe magazine. He recently showed that an agent can be applied for up to 40 minutes with no apparent changes to the tissue.

The next big step will be to transfer the studies from animal to human subjects. Although application of clearing agent to the skin of humans has been studied, Tuchin says that few have led to quantifiable results. He plans to trial injection-based studies on humans in the near future, once he is sure that the chemical agent is sufficiently sterile. Welch's group will examine the safety of the agents at useful concentrations, and determine the exact effects of the chemical agents on tissue.

Story courtesy of Opto and Laser Europe magazine