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A chip to support three-dimensional tissue culture

Is your liver damaged? No problem, we’ll grow you a new one. Isn’t this what we hope “regenerative medicine” could do for us? Nevertheless, researchers still do not have the means to grow organs in Petri dishes. Although it is possible to culture many cell types in vitro, including in co-culture with other cell types, the correct social and spatial behaviour of these cells depends on complex conditions that can only be simulated with difficulty by current cell culture systems. The KITChip, developed by researchers from the Karlsruhe Institute of Technology (KIT), improves the three-dimensional self-organisation of cells by enabling the active flow and circulation of the cell culture media. Using smart microtechnical methods, the scientists are able to adapt the microenvironment of cell types to their specific requirements, which leads to specific cellular behaviour and development. At present, the researchers are using the KITChip to simulate stem cell niches for basic research purposes.

Two KITChip variants under the laser scanning microscope r- (left) and f-KITChips (right) inoculated with HepG2 haematoma cells (staining: Syto 16). Top row: top view; below: cross section. © Dr. Eric Gottwald

Stem cells are the future of regenerative medicine. Because of their ability to generate almost any type of body cell, researchers anticipate that stem cells will be used in the future to repair damaged organs. The knowledge and understanding of appropriate culture conditions will soon enable tissue engineers to grow parts of the liver, the kidneys or spleen in a Petri dish and transplant the cultivated organs into the body. At least, this is what researchers are hoping. However, the truth of the matter is far more complicated. Body cells are normally embedded in a complex environment consisting of cells and extracellular matrix, the defining feature of connective tissue in animals and humans. The extracellular matrix, consisting of proteins and sugars, provides structural support to the cells in addition to performing many other important signalling functions. This matrix is also important for the development of tissue-specific structures: where should blood vessels grow? In which direction should specific cell types migrate to establish a functional whole? "Our KITChip with its design variants supports the formation of three-dimensional tissue structures," said Dr. Eric Gottwald from the Institute of Biological Interfaces 1 (IBG 1) of the Karlsruhe Institute of Technology (KIT). "We are able to improve the cultivation environment for many cell types by culturing the cells in an environment that closely resembles the organ of origin."

Suitable environmental architectures

Three KITChip variants with different types of microcavities © Dr. Eric Gottwald

The KITChip consists of an inconspicuous polymer plate whose external measurements are 20 mm x 20 mm and that has a central grid-like microstructured area consisting of 500 to 1,156 microcontainers that can harbour up to 10,000 cells each. The containers can vary in shape (cubic or spherical cavities in the polymer plate). The foil-based KITChip variant (f-3D-KITChip) has around 1 million pores per square centimetre, which enables the active flow and circulation of the cell culture media. The KITChips can be integrated into bioreactors that are adapted to specific applications. This peripheral infrastructure consists of a tube and pump system to generate a closed cycle through which the medium circulates. If necessary, this can prevent a gradient from forming above the tissue. On the other hand, a gradient can be established by changing the flow and circulation of the media.

But how do Gottwald and his team manage to culture the cells in an environment resembling that of the organ of origin? “In contrast to other three-dimensional culture systems, it is not only possible to actively supply our 3D cultures with nutrients, we can also modify the surfaces of the polymers used in order to establish suitable stem cell niches,” Gottwald explains. For example, Gottwald and his team can coat porous film sheets with collagen, a polypeptide that keeps our body cells together. Recently, the researchers also managed to specifically modify the polymer surfaces. They used UV radiation to insert hydroxyl groups into polymer chains close to the surface of the polymer. These hydroxyl groups improve the cells’ ability to adhere to the polymer surfaces. However, it is difficult to specifically apply such complex structures to the polymer surface, as they are only a few micrometres in size. One of Gottwald’s colleagues, Dr. Stefan Giselbrecht, has developed a method to produce functionalised microstructures from thin polymer films known as substrate modification and replication by thermoforming (SMART). The technology consists of three steps: preprocessing, microtechnical thermoforming and postprocessing. During the thermoforming process, the polymer film can be locally modified whilst preserving the patterned three-dimensional modifications.

Targeted cultivation through surface modifications: the photo on the left shows a non-modified KITChip matrix, whilst the one on the right shows the specific modification of the matrix structure.
Targeted cultivation through surface modifications: the photo on the left shows a non-modified KITChip matrix, whilst the one on the right shows the specific modification of the matrix structure. © Dr. Eric Gottwald

A niche for stem cells?

The KITChip has great future potential for the field of regenerative medicine. “The cells recognise the surface structures and settle in their environment,” said Gottwald. The Karlsruhe researchers have worked on numerous projects aimed at producing tissue that is very similar to the organ of origin (e.g., liver tissue). Their investigations have also shown that the KITChip platform leads to far better results than Petri dishes. “It would be great if we could produce the chips from bioresorbable polymers and use them directly as transplants,” said Gottwald.

“At the moment we are working on the development of a niche that is similar to the stem cells’ natural niche,” Gottwald explains. How can different types of cells be grown on the polymer to control the development of stem cells in an artificial cell complex? This is still pure basic research that is being carried out by interdisciplinary research groups involving biologists, engineers, physicists and chemists at the KIT. However, it is likely that these basic research activities will be of great benefit for regenerative medicine applications at some time in the future.

Further information:
Dr. Eric Gottwald 
Head of the Institute of Biological Interfaces 1 (IBG 1)
Hermann-von-Helmholtz-Platz 1
76344 Eggenstein-Leopoldshafen
Tel.: +49 (0)7247-822504
Fax.: +49 (0)7247-825546
E-mail: eric.gottwald(at)kit.edu

Website address: https://www.gesundheitsindustrie-bw.de/en/article/news/a-chip-to-support-three-dimensional-tissue-culture