Sustainable and responsible animal food production
Single-cell technology increases animal welfare
Lab animals are essential to be able to learn more about how an organ functions, yet the future may hold and alternative: organoids. These mini organs should be able to replace lab animals. The organoids need to be optimised first though, and single-cell technology, in which the genes of one single cell are deciphered, is helping to achieve that.
How do a fish’s intestines absorb nutrients and what happens in the intestines of a pig if it becomes infected, for example with a virus? If researchers want to investigate such topics, they need lab animals. If we want to use fewer lab animals in the future, we need to look for alternatives.
According to Animal Breeding & Genomics professor Martien Groenen, organoids could be the answer. Organoids are artificially grown, smaller, simplified versions of the organ, created by using stem cells that are cultivated in the lab. Groenen works with Ole Madsen, an associate professor of Animal Breeding and Genomics, on characterising organoids, including those for pigs and fish. By figuring out the make-up of the mini organs, they can help other groups within WUR develop better organoids.
“Intestinal organoids can let us study the effect of feed on the intestines, for example. We can also see whether animals are susceptible to certain bacteria or viruses without having to do tests on the animals themselves. This means we will eventually need fewer lab animals.” Organoids make good research models, but they are not identical to the original organ tissue. Madsen: “Ideally, of course, you would want the cultured tissue to be just as complex and complete as the intestinal tissue. But it will always be an artificial system, an approximation to the real world. So if you do research using organoids as a substitute for organs, it’s important to know exactly how they differ from real organs. That helps us to interpret the results better.” Groenen adds: “We compare the mini organs against the original tissue to see what is missing. Ultimately, the aim is to cultivate organoids that resemble the natural tissue even more closely.”
‘Ideally, you want the cultured tissue to be just as complex and complete as the intestinal tissue’
To find an answer to this question, Madsen and Groenen are using single-cell genomics, or single-cell technology. This is a technique that lets you see at the cell level which genes are active and which genes are switched off.
To this end, the tissue cells are first separated out, usually by applying enzymes that break down the protein links between the cells. Then the DNA (or RNA) is extracted from the cells and replicated to produce sufficient genetic material for analysis. In single-cell technology, this replication involves giving all the DNA or RNA from one individual cell a special code (address) so that it is possible to see in the analysis which DNA or RNA came from that cell.
Madsen: “If we do this with intestinal organoids, we see that the three thousand cells or so can be divided into five or six groups in which the same genes are switched on or off. In addition, the intestinal organoid may contain a few rare cells, which can be identified with single-cell technology. That gives a much better understanding of the composition and function of the tissue. We also see that not all the cell types found in actual intestinal tissue are present in the organoids. For example, this technique shows us that the immune cells that are so important in real intestines are missing in the mini intestines.”
‘We compare the mini organs against the original tissue to see how we can make a better replica of the organ’
Immune cells act like guards, subjecting everything that comes through the intestines to a selection process: is this an undesirable bacterium or a useful protein? Useful substances are allowed through, random samples are taken of some other substances to give the immune system information, while other substances ‘feed’ the bacteria and yeasts that form the microbiome. Immune cells are also crucial for communication with the microbiome.
“Therefore it is essential that immune cells are added to the mini intestines as those cells are such an important part of the organ,” explains Madsen. “Immune cells constantly patrol the surroundings on the lookout for intruders and focus on combating infections or removing dead cells.
A complete model of the intestines that include immune cells would give a much better picture of the influence of diet on the intestines’ functioning.” The Cell Biology and Immunology chair group is now working on cultivating intestine cells and immune cells at the same time. But it is quite a challenge to create an organoid that includes both intestine cells and immune cells. Madsen: “Different signal molecules are required during the differentiation, so the immune cells have to be prepared separately and then combined with the other intestinal cells at just the right moment. To make that process go smoothly, further special signal molecules are needed that tell the immune cells where they need to be to get them to migrate to the right point in the organoid tissue.”
‘How did evolution lead to the functional differences between species? We can study that on a cellular level’
This is advanced research, and that means steep costs. At present, the costs can still be an obstacle. Groenen: “In a follow-up project, we want to investigate how we can reduce the costs of this kind of research. We think this is important for the future, because we have noted interest among animal breeding companies. They need to be able to detect genetic differences in susceptibility to bacteria and viruses and select for that within a population. We could also help animal feed companies develop feed that is even healthier.”
“At the moment, we are still focusing on immune cells and organoids of pigs’ intestines. But the expectation is that we will be able to use the technique for organoids of other creatures’ organs, such as fish.
We are very interested in the mechanisms of different animals: how evolution led to the functional differences between species. That is the miracle of nature, which we will be able to study on a cellular level.”
Madsen: “What is it that makes one animal able to adapt to conditions while another animal can’t? If we study the influence of the environment on the development of an animal, we end up with variations in the genome and variations in the genes that are switched on or off — epigenetics. We want to know how all those pieces of the puzzle fit together. But the more details we see, the more things we discover we don’t yet fully understand.”
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TEAM Richard Crooijmans, Alisha GM van Animal Breeding & Genomics
Researchers portrayed in this article: Martien Groenen, Ole Madsen with Alisha GM
MORE INFORMATION This project is part of the Next Level Animal Sciences (NLAS) innovation programme.
Participating researchers of Wageningen University & Research collaborate with various partners to develop new research methods and technologies within the field of animal sciences. NLAS consists of three research directions, namely sensor technology, complex cell systems and data and models.