The immortal liver cell
The liver of a modern high-producing cow has to do amazing things. It is mainly responsible for producing glucose which, in turn, is needed to produce large quantities of milk. This can overload the liver and prevent it from performing other metabolic activities. It can also lead to a fatty liver or ketosis. “One in two cows becomes ill immediately after giving birth to a calf and can only be used for milk production to a limited extent”, says Professor Marion Schmicke from the Institute of Agricultural and Nutritional Science. A dairy cow’s life expectancy could also be increased by at least two to three years if these illnesses were avoided.
That’s why it is very important for veterinary researchers to gain a better understanding of bovine liver – but this is highly complicated. “It’s not easy to cultivate bovine hepatocytes, also known as liver cells”, explains Schmicke. However, it would make research much easier from both an ethical and financial point of view. “Cows are big and therefore expensive to keep”, says the scientist. The results of experiments on other animals (e.g. rats and mice) are difficult to transfer to cattle, as their livers are under much less strain.
Cells survive for 30 days
Schmicke’s research group is therefore looking to develop a new cell line consisting of bovine liver cells (i.e. an immortal cell culture). This will then serve as a model for testing new drugs, researching diseases such as ketosis and for other purposes. To do this, cells of a certain tissue type – in this case hepatocytes – must be modified in such a way that they permanently survive while remaining as similar as possible to the original cells. The first step has been taken within the cooperative pan-European project, BovReg, which is being funded through the EU’s Horizon 2020 programme. “We’ve reached the stage where the removed liver cells can survive in the laboratory for 30 days”, says Schmicke. They are also much more closely related to the original liver cells than previous cultures of bovine liver cells. This has been achieved by optimising the culture medium in which the cells grow. “There are very few research groups around the world that can even produce such cell cultures”, explains the veterinarian. This is because hepatocytes are highly sensitive.
However, the cells have to continuously multiply to facilitate the cultivation of a cell line and ensure their permanent survival. “The body’s cells usually stop multiplying after a certain number of cell divisions”, says Schmicke. This is partly caused by so-called “telomeres”, which are DNA sequences at the end of chromosomes that become shorter with every cell division. However, some cell types contain telomerase, an enzyme that rebuilds the telomeres and ensures that things like bone marrow cells, stem cells and egg cells can multiply without loss. It is also active in most cancer cells, which is the cause of their abnormal proliferation.
A long road ahead
“So this telomerase has to be active for a cell line to permanently survive”, explains Schmicke. There are several ways of achieving this. For example, the cells can be treated with so-called “oncogenes”, which means artificially turning them into cancer cells. However, the scientist explains the disadvantages of this method: “A tumour cell is simply no longer a normal cell”. The aim is ultimately to activate the telomerase while modifying the cells as little as possible, as this is the only way for research into cell lines to provide useful information as to what happens in the corresponding cells of an entire organism.
Last year, Schmicke’s research group set up an S2 laboratory (i.e. a laboratory with a safety level of two), where genetic engineering can be carried out in compliance with the relevant safety regulations. This enables the hepatocytes to be modified in such a way that they produce telomerase. “We have been experimenting with bacterial plasmids”, says Schmicke. These are small, ring-shaped pieces of DNA that can be used to introduce foreign genetic information in cells. The plasmids can be designed specifically in such a way that only the desired changes take place in the cell. Although this process has worked, the telomerase introduced in the cells has remained inactive. “Now we’re going to experiment with artificially modified viruses”, says the scientist. Viruses are very good at introducing genetic information in cells and then reprogramming them for their own purposes. The viruses will be modified to stop them from multiplying and to make sure that they mainly introduce the required genes in the cells.
So, there is still a lot of work to be done before a cell line can be developed from the bovine liver cell culture, and regular tests have to be carried out to make sure the modified hepatocytes are as close as possible to the original. “But we’re on the right track”, says Schmicke. Despite the COVID restrictions, which have made it difficult for the research group to get their hands on fresh bovine livers, the first step is completed. “This is mainly thanks to our doctoral candidate, Sandra Andres, who has battled through the difficulties to achieve things that can sometimes take three times as long”, says Schmicke. Andres is now a veterinarian in Baden-Württemberg, where she continues to work with cattle. The work on the new cell line is now being continued by other members of the research group.