According to Cambridge researchers, roughly three-quarters of all stem cell lines grown from human skin cells have DNA damage brought on by elements like UV light. They contend that whole genome sequencing is crucial for determining whether or not cell lines are viable.
A unique class of cells called stem cells can be instructed to develop into practically any form of cell in the body. Currently, they have employed in research on the early phases of the embryo as well as the development of organs.
Researchers are increasingly using stem cells to create novel therapeutics known as cell-based therapies. Other potential uses include engineering stem cells to develop into nerve cells in illnesses like Parkinson’s to replace those lost to neurodegeneration.
Stem cells were originally created from embryonic tissue, but now adult skin cells can also be used to create stem cells. These so-called induced pluripotent stem cells (iPSCs) may now be produced from a variety of tissues, including blood, whose popularity is growing as a result of how simple it is to obtain.
However, there is an issue with stem cell lines made from both skin cells and blood, according to researchers at the University of Cambridge and the Wellcome Sanger Institute. When scientists closely investigated the stem cell lines’ genomes, they discovered that about 75 percent of them had significant DNA damage that could make it difficult for them to be used for research and, more importantly, cell-based therapeutics. Their findings, which are now published in Nature Genetics, represent the broadest genomic analysis of iPSCs to date.
Three billion pairs of nucleotides, or molecules denoted by the letters A, C, G, and T, make up DNA. Damage to our DNA over time, such as that caused by ultraviolet light, can result in mutations. For instance, a letter C might become a letter T.
Our DNA contains “fingerprints” that can be used to identify the cause of this harm. As these mutations multiply, they may significantly affect how cells operate and, in certain cases, result in tumors.
In recent years we have been finding out more and more about how even our healthy cells carry many mutations and therefore it is not a realistic aim to produce stem cell lines with zero mutations. The goal should be to know as much as possible about the nature and extent of the DNA damage to make informed choices about the ultimate use of these stem cell lines. If a line is to be used for cell-based therapies in patients for example, then we need to understand more about the implications of these mutations so that both clinicians and patients are better informed of the risks involved in the treatment.
Dr. Foad Rouhani
Dr. Foad Rouhani, who carried out the work while at the University of Cambridge and the Wellcome Sanger Institute, said:
“We noticed that some of the iPS cells that we were generating looked really different from each other, even when they were derived from the same patient and derived in the same experiment. The most striking thing was that pairs of iPS cells would have a vastly different genetic landscape one line would have minimal damage and the other would have a level of mutations more commonly seen in tumors. One possible reason for this could be that a cell on the surface of the skin is likely to have greater exposure to sunlight than a cell below the surface and therefore eventually may lead to iPS cells with greater levels of genomic damage.”
When the whole DNA of stem cell lines from several cohorts, including the HipSci cohort at the Wellcome Sanger Institute, was examined by the researchers using a popular method called whole genome sequencing, they found that up to 72% of the lines displayed significant UV damage.
Professor Serena Nik-Zainal from the Department of Medical Genetics at the University of Cambridge said: “Almost three-quarters of the cell lines had UV damage. Some samples had an enormous amount of mutations sometimes more than we find in tumors. We were all hugely surprised to learn this, given that most of these lines were derived from skin biopsies of healthy people.”
They made the decision to shift their focus to cell lines not produced from the skin and concentrated on iPSCs obtained from blood as these are growing in popularity due to the simplicity of getting blood samples. They discovered that although these iPSCs from blood also harbored mutations, the level of mutations was lower than in iPS cells from the skin, and there was no UV damage.
However, a gene termed BCOR, a crucial gene in blood malignancies, was mutated in about 25% of the population. They differentiated the iPSCs into neurons and followed their development to determine whether these BCOR alterations had any functional effects.
Dr. Rouhani said: “What we saw was that there were problems in generating neurons from iPSCs that have BCOR mutations they had a tendency to favor other cell types instead. This is a significant finding, particularly if one is intending to use those lines for neurological research.”
The process of cell culture appears to increase the frequency of these mutations, which may have implications for other researchers working with cells in culture. When they looked at the blood samples, they found that the patient did not have the BCOR mutations; rather, they were not present in the patient.
Typically, researchers analyze the chromosomes of their cell lines for issues, such as whether they have the necessary 23 pairs of chromosomes. This, however, would not be thorough enough to catch the potentially serious issues that this recent study has found.
Importantly, scientists and doctors would not be aware of the underlying damage that exists in the cell lines they are dealing with without closely examining the genomes of these stem cells.
“The DNA damage that we saw was at a nucleotide level,” says Professor Nik-Zainal. “If you think of the human genome as like a book, most researchers would check the number of chapters and be satisfied that there were none missing. But what we saw was that even with the correct number of chapters in place, lots of the words were garbled.”
Fortunately, says Professor Nik-Zainal, there is a way around the problem: using whole genome sequencing to look in detail for the errors at the outset.
“The cost of whole genome sequencing has dropped dramatically in recent years to around £500 per sample, though it’s the analysis and interpretation that’s the hardest bit. If a research question involves cell lines and cellular models, and particularly if we’re going to introduce these lines back into patients, we may have to consider sequencing the genomes of these lines to understand what we are dealing with and get a sense of whether they are suitable for use.”
Dr. Rouhani adds: “In recent years we have been finding out more and more about how even our healthy cells carry many mutations and therefore it is not a realistic aim to produce stem cell lines with zero mutations. The goal should be to know as much as possible about the nature and extent of the DNA damage to make informed choices about the ultimate use of these stem cell lines.”
“If a line is to be used for cell-based therapies in patients for example, then we need to understand more about the implications of these mutations so that both clinicians and patients are better informed of the risks involved in the treatment.”
The study was supported by the UK Regenerative Medicine Platform and the NIHR Cambridge Biomedical Research Center, with funding from Cancer Research UK, the Medical Research Council, and Wellcome.
