Piglets for transplantologists
Endogenous retroviruses were completely removed from pig DNA
With the help of CRISPR genome editing technology, scientists were able to cut out endogenous retroviruses from the pig genome – genetic traces of viruses that once embedded their DNA in pig DNA. As a result of the experiment, healthy piglets were born. The achieved result, according to many experts, will greatly facilitate the cultivation of pig organs suitable for human transplantation and make them safer.
Once upon a time, these viruses penetrated into cells, created a fragment of a DNA chain based on their RNA, which they embedded into the cellular genome to create their copies. But then the body coped with the viral infection, and the DNA built by the virus turned into a non-functioning fragment of the genome, transmitted from generation to generation. Humans also have their own endogenous retroviruses, they make up up to 8% in our genome. Some of them even began to play a role in regulating the work of other human genes.
There are more than 50 endogenous pig retroviruses (the exact number depends on the breed), divided into three groups: PERV-A, PERV-B and PERV-C. Experiments show that some retroviruses from the PERV-A and PERV-B groups can infect human cells in culture, which means a potential danger of infection during transplantation of an organ grown in a pig's body to a human. In addition, the activity of endogenous retroviruses, as the study shows, significantly reduces the chances of normal engraftment of the transplanted organ. You can read more about this in the essay "Pig's Heart".
Geneticists from Harvard University George Church (George Church) and Luhan Yang (Luhan Yang), one of the pioneers of the CRISPR method, founded in 2015 the company eGenesis, specializing in obtaining genetically engineered organs for transplantation. In the same year, they were able to modify 62 pig genes related to endogenous retroviruses. But for this project, the researchers used a line of "immortal" pig kidney cells capable of living and dividing indefinitely in cell culture.
Now it's the turn of genetically normal cells taken directly from a live pig. In the new work, eGenesis collaborators, together with Danish and Chinese scientists, applied the CRISPR system to cells obtained from the connective tissue of pig embryos. After genome editing, normal cell growth was disrupted. Perhaps, according to Luhan Yang, DNA damage prompted them to stop dividing or started the process of self-destruction. But by acting on cells with substances that stimulated growth and blocked the gene responsible for suppressing growth, scientists were eventually able to multiply cells that do not contain any endogenous retroviruses in cell culture.
The researchers then applied a standard cloning procedure. They took cell nuclei containing DNA and placed them in pig eggs. The resulting embryos were implanted into the uterus of the surrogate mother. "Before our study, there was a huge scientific uncertainty as to whether a pig [born after such an edit] would be viable," says Luhan Yang. As it turned out, the survival rate of embryos in the study does not differ from their survival rate during normal cloning (about one embryo out of a hundred implanted). A tissue check of 37 born piglets showed that there are really no endogenous retroviruses in their genomes.
The study is published in the journal Science.
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29.08.2017