Dr. Lothar Koch: Organs from the printer – Three-dimensional “cell printing” for producing biological tissue

3D printers for private use can be bought for less than 1,000 euros. These are able to print three-dimensional structures from thermoplastic synthetics. There are also 3D printers for other materials. Printing techniques like these have also been tested for medical use for several years now.

 

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Printed pattern with fluorescent cells. The diameter of the printed cell-sol-drop is a few 10s to several hundred micrometres. This corresponds to volumes of a few picolitres to several nanolitres.

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After printing, the cells do not necessarily stay in one place. Depending on the environment, they move to some extent (here after 2 days). This must be considered when complex cell systems or biological tissues are printed.

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Time-resolved depiction of the laser-based printing process. In the first microsecond a steam bubble builds and collapses again. It accelerates the cell-sol-mixture, which afterwards continues flowing as a thin spurt.

A new, interesting area is the printing of individualised implants or the printing of supporting structures (scaffolds), which can afterwards be used as bone or cartilage replacements when they are populated by cells. But efforts are also being made to print the cells themselves, in order to produce organic tissue. In the future, perhaps, it might be possible to produce complete replace­ment organs. This would not only counter the lack of organ donations, but rejection reactions could be prevented by using the patients’ own cells and the life-long intake of immunosuppressives, which follows from normal trans­plantations, would no longer be necessary.

Before printed tissues and organs can be transplanted in patients, there are also several possible usages for printed cell structures in research. A broad understanding of inter­­­­actions between different cells and their environment is necessary to be able to produce complex tissues and to conduct (stem) cell-based therapies. Conventional cell studies in two-dimensional cell structures (Petri dishes, in vitro) are not suitable for simulating the complex inter­actions in three-dimensional tissues and in cell microen­vi­ronments like those in the body (in vivo). Cells react fundamentally differently in 3D.

Therefore, printed 3D cell patterns can fill the gap be­­tween conventional cell cultures in vitro and animal models in vivo. Innovative 3D cell models could lead to new insights into cell behaviour, tissue functions and regeneration and effects of bioactive substances and pharma­ceuticals. In the long run, by using human cells they could exceed animal models in relevance for medicine and even replace them.

Different printing techniques are being tested for the printing of living cells. Generally no “naked” cells are being printed, instead these are mixed/suspended into a medium. Usually water-based gels (hydrogels), respec­­tively their non-gelatinised precursors, called sol, are used for this. These have to fulfil multiple functions: they need to be suitable for the printing process, whereby viscosity plays an important role. As a so-called extra-­cellular matrix, they should provide stability and structure to the printed tissue piece after printing, wherefore the sol can be gelatinised through the printing of another, corresponding chemical substance. They must provide the cells with a life-sustaining, nutritious environment. They should not unwantedly influence the be­­haviour of the cells after printing. Of special interest are those gels that also exist naturally in the human body, such as collagen, fibrin, and hyaluronic acid. However, the gel needs to be chosen correspondingly to the cell type or the tissue that will be printed.

Modified ink-jet printer are used for printing. The ink is replaced by a cell suspension. These produce a high resolution. However, only cell-sol-mixtures with low vis­­cosity and low cell density can be used, as otherwise the nozzles become clogged. The cell density in natural tis­sues, like for example the upper layer of skin (epidermis), cannot be achieved. On the other hand, extrusion systems are used. There, the cell-sol-mixture is squeezed through nozzles with an inner diameter of 50 to 1,000 micrometres, similar to a syringe or a pipette. The smaller nozzle diameters have similar disadvantages as the ink-­jet printers. With bigger diameters, the print resolution suffers, however, higher cell densities and gels with a higher viscosity can be used.

A nozzle-free, laser-based technology, which permits combining a high cell density and a high viscosity with a high resolution is being tested at the Laser Zentrum Hannover in the nanotechnology department. It is already used in a modified version for printing conducting paths onto, for example, solar cells. In the biomedical application, small drops of the cell-sol-mixture with a volume in the picolitre range (few 10 micrometres in diameter) are “shot out off” a layer of sol via laser. Now, a freely selectable pattern can be printed drop by drop and layer by layer, including a three-dimensional model.

Different research groups have investigated thoroughly whether the printing process damages or influences the cells. It was ascertained that, if suitable parameters are chosen, almost 100 per cent of the cells survive the print­ing process and are not altered. Stem cells are also not influenced in their differentiation behaviour. Therefore stem cells were already printed into defined patterns and differentiated into bones or cartilage.

In cooperation with Medizinische Hochschule Han­nover (Hannover Medical School, MHH) we already printed three-dimensional cell models to examine the interaction between blood vessel cells (endothelial cells and stem cells). In another cooperation with MHH, we printed skin, which was implanted on mice. It grew together with the mice’s skin and made up a structure similar to the mice’s skin. This printed skin is still lacking several func­­tions of natural skin, but further-developed with additional cell types it could, for example, replace animal ex­­periments for cosmetics tests.

Currently, many groups around the world are working on making 3D printing technology usable for medicine. Decades will pass, however, before a printed replacement heart made of living cells for patients becomes available. However, 3D printing already opens up many new and exciting fields of research.

 

The author studied mathematics and physics at the Leibniz University Hannover. From 2003 to 2007, he worked as a research assistant at the Institut für Quantenoptik at the Leibniz University Hannover. In 2007, he earned his doctor of science. From June 2007 on, he was project manager at the Laser Zentrum Hannover and in 2011 he took over the management of the Biofabrication Group there.