One step closer to artificial rhino eggs

Date:
March 10, 2022
Source:
Max Delbru"ck Center for Molecular Medicine in the Helmholtz
Association
Summary:
To prevent the extinction of the northern white rhino, researchers
are attempting to create artificial egg cells from stem cells. A
team has now revealed that they are one step closer to achieving
this goal.



FULL STORY ==========================================================================
To prevent the extinction of the northern white rhino, the international consortium BioRescue is attempting to create artificial egg cells from
stem cells. A team led by MDC's Sebastian Diecke and Micha Drukker of
Leiden University has now revealed in Scientific Reports that they are
one step closer to achieving this goal.


==========================================================================
Fatu and Najin are the last two northern white rhinos in the world --
and they are both female. As a result, this subspecies can no longer
reproduce naturally, and extinction seems inevitable. However, the
BioRescue consortium is working against the clock to ensure that the
northern white rhino does not disappear from our planet forever. The researchers are pursuing a two-pronged approach: First, they are
developing advanced assisted reproduction techniques; second, they want
to use skin cells taken from the northern white rhinoceros to create
induced pluripotent stem (iPS) cells in the lab, which can eventually
develop into immature egg cells, or oocytes. The team from the Max
Delbru"ck Center for Molecular Medicine in the Helmholtz Association
(MDC), together with partners in Munich, the Netherlands and Japan,
has now taken a major step towards this goal. In the journal Scientific Reports, they reveal that they have managed to obtain and conduct in-depth studies on pluripotent rhino stem cells. "Our paper sheds new light on pluripotency -- the ability of stem cells to differentiate into all cells
of the body," says lead author Dr. Vera Zywitza of the Pluripotent Stem
Cells Platform at the MDC, which is led by Dr.

Sebastian Diecke. "It therefore represents an important milestone on the
road to artificially generated rhino oocytes." The BioRescue project
has received EUR4 million in funding from the German Federal Ministry
of Education and Research (BMBF). The international consortium, which
includes the MDC, is led by the Leibniz Institute for Zoo and Wildlife
Research (IZW) and cooperates with many more partners, including Helmholtz Zentrum Mu"nchen in Germany.

The fine art of cell engineering iPS cells in a petri dish have
the potential to develop into any cells of the body -- including the
primordial germ cells that the BioRescue scientists want to cultivate. The researchers are working closely with the lab of Professor Katsuhiko
Hayashi, a Japanese stem cell researcher at Kyushu University. In
2016, Hayashi succeeded in generating egg cells from the skin of mice, artificially fertilizing these cells, and implanting them in females. The
mice conceived by this method were born healthy and fertile.

Stem cell researcher Professor Micha Drukker and his teams at Helmholtz
Zentrum Mu"nchen and at Leiden Academic Centre for Drug Reserach at
Leiden University used a process known as episomal reprogramming to successfully produce northern white rhino iPS cells. This involved
the researchers introducing foreign DNA molecules called "plasmids"
into skin cells they had obtained. These plasmids contained genes to
reprogram the skin cells into iPS cells. The rhino stem cells generated
in this way are remarkably similar to their human equivalent.

"Viewed under the microscope, they are barely distinguishable from human
iPS cells," says Drukker, adding: "They also respond very similarly to
external influences." A promising start for germline cell cultivation


==========================================================================
iPS cells have different states: they can be nai"ve -- the "ground
state" of pluripotency -- or primed. Cells in the latter state are
thought to have reached a slightly more advanced stage of embryonic development. Experiments with stem cells generated from mice show that
they are particularly good at producing germline cells when converting
from the primed to the nai"ve-like state. However, when the scientists
first attempted to convert the rhino cells to a nai"ve-like state,
the cells died. The researchers therefore introduced a gene into the
rhino cells that prevents cell death -- and with this, they successfully obtained nai"ve iPS cells. "We have characterized the cells in detail by,
among other things, analyzing transcriptome data," Zywitza explains.

"The successful conversion to a nai"ve-like state of pluripotency is a promising starting point for generating germline cells." Nevertheless,
Zywitza and her colleagues cannot yet move onto the next stage.

"The iPS cells we have cultivated contain persistent foreign genetic
material - - namely, the reprogramming factors and the gene that prevents
cell death," Zywitza explains. "This means we can't use them to make germ cells, as there is a risk these would be pathologically altered." But
these cells are still extremely useful for studying rhino stem cells in
general and gaining a better understanding of their different states in specific. With their help, scientists can explore the molecular mechanisms
that take place in stem cells.

"For example, we can study why the gestation period of a rhinoceros
is 16 months whereas that of a mouse is only 21 days, or how organs
develop in different species," the scientist explains. This teaches us
a lot about evolution." Ovarian tissue is needed too In the meantime,
Diecke's team has generated further iPS cells. They used RNA viruses
instead of plasmids to introduce the reprogramming factors. These new
iPS cells do not contain anything that does not belong there. Now the scientists are trying to produce primordial germ cells from them.

And that's not all: primordial germ cells only mature into egg cells
when they are surrounded by ovarian tissue. It is nearly impossible
to obtain such tissue from living or deceased rhinos. "So we have to
create not only primordial germ cells but also ovarian tissue," Zywitza explains. The Berlin-based scientists are working closely with Hayashi
to achieve this. Last year he successfully cultivated ovarian tissue
from mouse stem cells.



==========================================================================
14 embryos created so far using assisted reproduction Meanwhile, progress
is also being made with assisted reproduction. In January of this year, scientists at IZW, in collaboration with the Kenya Wildlife Service,
the Wildlife Research and Training Institute, the Dv?r Kra'love' Safari
Park, and Ol Pejeta Conservancy, collected immature egg cells called
oocytes from Fatu. These were matured at Avantea's laboratories in Italy
and inseminated with thawed sperm from a deceased bull. There are now
a total of 14 northern white rhino embryos, which are stored in liquid
nitrogen at minus 196 degrees Celsius. In the near future the embryos
will be implanted into southern white rhino surrogates, with the aim of creating a healthy northern white rhino calf.

The production of 14 embryos represents a great success in reproductive biology, but this is not a large amount if the goal is to rebuild the
northern white population to a self-sustaining level. "Najin and Fatu
are also closely related and their genetic makeup is largely identical,"
says Professor Thomas Hildebrandt of IZW, who leads the BioRescue research consortium. "Due to age and reproductive tract issues, we were unable to collect any oocytes from Najin that could be developed into embryos, so
all 14 embryos are from Fatu. We therefore urgently need a complementary strategy for creating gametes -- eggs and sperm -- from significantly
more individuals." Protecting the world's species -- before it's too late "Generating functional eggs of the northern white rhinoceros would be the crowning achievement of our research," Diecke says. This approach could
serve as a model for other endangered species. If reproduction from stem
cells works, the technique could be used to revive many more threatened
or already extinct species. More than 10,000 living cell cultures from
more than 1,000 endangered species are stored in the Frozen Zoo at the
Arnold and Mabel Beckman Center for Conservation Research in San Diego
and in IZW's biobank in Berlin. "This invaluable resource could be used
to bring back species from the brink of extinction," Diecke says. The
northern white rhino would then be just the beginning -- although he
adds that he "would rather we never had to use our technique and did
more to preserve species before it was too late." But for Zywitza,
one thing is certain: If a northern white rhino is born someday thanks
to stem cell technologies, she would love to meet it.


========================================================================== Story Source: Materials provided by Max_Delbru"ck_Center_for_Molecular_Medicine_in_the
Helmholtz_Association. Original written by Jana Ehrhardt-Joswig. Note:
Content may be edited for style and length.


========================================================================== Related Multimedia:
* Rhinos_and_stem_cells ========================================================================== Journal Reference:
1. Vera Zywitza, Ejona Rusha, Dmitry Shaposhnikov, Jorge Ruiz-Orera,
Narasimha Telugu, Valentyna Rishko, Masafumi Hayashi, Geert
Michel, Lars Wittler, Jan Stejskal, Susanne Holtze, Frank
Go"ritz, Robert Hermes, Jichang Wang, Zsuzsanna Izsva'k, Silvia
Colleoni, Giovanna Lazzari, Cesare Galli, Thomas B. Hildebrandt,
Katsuhiko Hayashi, Sebastian Diecke, Micha Drukker. Nai"ve-like
pluripotency to pave the way for saving the northern white
rhinoceros from extinction. Scientific Reports, 2022; 12 (1) DOI:
10.1038/s41598-022-07059-w ==========================================================================

Link to news story: https://www.sciencedaily.com/releases/2022/03/220310115102.htm

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