Although only two females remain, the end of the northern white rhino is not yet sealed: researchers report successes in the goal of conserving the species using stem cell technology. They have succeeded in generating so-called pluripotent stem cells from frozen skin cells of a specimen that has already died. The scientists hope that they could now be used to generate rhinoceros ova.
Habitat loss and above all the coveted horn were the fatalities of the northern white rhinoceros (Ceratotherium simum cottoni). Then, in 2018, came the news that seemed to finally seal the fate of the species: the last bull had died. Only the two rhino ladies Fatu and Najin were left. As a result, natural reproduction possibilities were no longer available. Nevertheless, the friends of the charismatic pachyderm did not want to give up: For the continued existence of the species, all registers of reproductive technology are being pulled out within the framework of the international BioRescue consortium.
The scientists are pursuing two strategies: methods of assisted reproduction are intended to ensure offspring and they also want to generate stem cells and ultimately egg cells in the laboratory from skin cells of the northern white rhinoceros. As far as assisted reproduction is concerned, the team has already come a long way: egg cells were taken from the rhinoceros Fatu and fertilized in the laboratory with the thawed sperm of the last bull. In the meantime, 14 embryos have emerged that are now being stored in liquid nitrogen. They will soon be implanted in southern white rhino surrogate mothers to provide offspring.
A second approach is necessary
However, this is not enough to provide a basis for the survival of the species. “Najin and Fatu are too closely related and their genes are partly identical,” says BioRescue project manager Thomas Hildebrandt from the Leibniz Institute for Zoo and Wildlife Research. “Due to her age and impairments in the reproductive tract, we were also unable to obtain any oocytes from Najin from which embryos could be successfully created. All 14 embryos are therefore from Fatu. We therefore urgently need a complementary strategy to generate gametes – i.e. egg cells and sperm – from significantly more individuals.”
The team is therefore also pursuing the stem cell strategy. The team led by Vera Zywitza from the Max Delbrück Center for Molecular Medicine in Berlin (MDC) has now taken a big step forward. As they explain, special units are required for the production of egg cells: so-called induced pluripotent stem cells (iPS cells), which can theoretically be obtained from any cell in the body. In developing their method, Zywitza and her colleagues are working closely with researchers who have already been successful in mice: in 2016, the team led by Katsuhiko Hayashi from the Japanese Kyushu University succeeded in generating egg cells from rodent skin cells, artificially inseminating them and giving them female eggs to plant mice. This resulted in healthy and fertile offspring.
However, a rhino is not a mouse – the transfer of the method proved to be difficult. Nevertheless, the BioRescue team has now succeeded in producing rhinoceros iPS cells from skin. The starting material came from the rhino cow Nabire, who died in a zoo in 2015. As the researchers report, they were successful with the technique of so-called episomal reprogramming. In doing so, they first introduced additional genetic information into the skin cells via special DNA carriers – so-called plasmids. They mediated functions that were ultimately able to transform skin cells into so-called naïve iPS cells. “Successful conversion to the naïve state of pluripotency provided a promising starting point for generating germline cells,” says Zywitza.
chance of survival
The cells generated by the plasmid method are well suited to researching the stem cells of the rhino and to better understand their different states. However, they cannot be used to generate germ cells because they still contain the disruptive foreign genetic material, the scientists explain. However, as the MDC reports, the team has now also succeeded in generating iPS cells using an alternative method. The reprogramming factors were not introduced by plasmids, but with the help of RNA viruses. These new iPS cells then no longer contain anything that doesn’t belong, the statement says. This material can now be used to produce progenitor cells of egg cells.
The researchers report that another component that is necessary for the success of the project could also arise from the iPS cells: ovarian tissue. As they explain, progenitor cells only mature into egg cells if they are surrounded by this material. “So we have to create progenitor cells as well as ovarian tissue,” says Zywitza. The BioRescue team is also in close contact with Katsuhiko Hayashi. Because he has already successfully generated ovarian tissue from mouse stem cells.
“Functional egg cells of the northern white rhino – that would be the culmination of our research work,” says Sebastian Diecke from the Max Delbrück Center. According to him, the successes could also serve as a model: If reproduction from stem cells in rhinos is successful, other endangered species or species that have already been exterminated by humans could also be revived in this way. Starting material is often readily available: Cell cultures of more than 1,000 endangered species are stored in the Frozen Zoo at the Beckman Center for Conservation Research in San Diego and in the biobank of the Institute for Wildlife Research in Berlin. In conclusion, however, Diecke emphasizes: “However, I would like it best if our approach never had to be used and more was done for species conservation before it is too late,” says the scientist.