UC Berkeley Berkeley Scientific Journal

Home , Cryopreservation, Cryoprotectant

Title Frozen in Time

Permalink https://escholarship.org/uc/item/8rh8n3w6

Journal Berkeley Scientific Journal, 21(1)

ISSN 1097-0967

Author Tran, Phuong

Publication Date 2016

DOI 10.5070/BS3211033749

Undergraduate

eScholarship.org Powered by the California Digital Library University of California FROZEN IN TIME

BY PHUONG TRAN

“WITH THE USE OF CRYOPRESERVATION, DE-EXTINCTION MAY BECOME A POSSIBILITY IN THE NEAR FUTURE.”

hould we bring back extinct species? and thaw them in the future for in vitro breeding. Unfortunately, inbreeding holds SMany endangered species in the wild fertilization (IVF). IVF is the process of potential problems such as: increased are facing the possibility of extinction. creating embryos from eggs and sperm reproductive failures leading to fewer of- However, thanks to the developments of by fertilizing them outside of the womb spring, genetically undesirable individuals, genetic technology, extinction does not and then transferring the embryo into and widespread damaging genes. However, have to be permanent. It is very possible to the recipient. Tus, scientists hope that this is not to say that inbreeding does not resurrect some of the species we once lost cryopreservation is the solution to saving occur naturally. Certain species repro- and even prevent endangered species from endangered species and potentially bring duce outside their breed on their own for ever becoming extinct. For many years, back extinct species. their own survival. Inbreeding is a double scientists have played around with the An endangered species is a group of edge sword. Te results of inbreeding can possibility of “de-extinction”1 and alterna- organisms that are at risk of becoming produce excellent quality animals. Howev- tive methods to repopulating endangered extinct for one or more of the following er, excessive inbreeding can make matters species. With the use of cryopreservation, reasons: destruction of their living envi- worse for the species. Tus, scientists have de-extinction may become a possibility in ronment, an increase in predators, and/or turned to cryopreservation for solutions. the near future. Cryopreservation is the unsustainable breeding due to low member Survival of some endangered species the process in which certain cells such as population. Over the years, scientists and heavily relies on “frozen zoos.”3 Te Fro- the eggs and sperm are preserved in very researchers have explored numerous tech- zen Zoo at San Diego’s Institute for Con- low temperatures and then extracted at niques in hopes of reintroducing endan- servation Research was founded in 1972 a later time when needed. By storing the gered and even extinct animals back into as a storage for skin-cell samples from rare cells in very low temperatures, it allows to the wild. Most endangered species are kept and endangered species. When the frst extend the lifetime of cells outside their in captivity to breed with the remaining samples were collected and frozen, genetic original hosts. In a way, cryopreservation members of their population. Some spe- technology was still in its new stages so allows us to freeze time and preserve the cies have even been forced to breed with it was not really known how the samples eggs and sperms of endangered species closely related species, also known as in- would be used. “Te Frozen Zoo was a

50 Berkeley Scientifc Journal | FALL 2016 wonderful idea. Tey just thought: rate to maintain the current population. ‘Well, something might happen, so we Many scientists predict that the Asian ele- should preserve some samples for the phant will become extinct within the next “Extinction future,” says Dr Jeanne Loring, who is few decades if their fertility rates continus leading the Scripps team working with to decline. frozen samples.3 “Tis is the frst time At the Hannover Zoo in Germany, does not have that there has been something that we thirty ejaculates were collected from six can do.”3 Cells preserved in frozen zoos Asian elephants and one African elephant, to be can be added to the gene pool, increas- a close living relative to the Asian ele- ing the chances of healthy reproduction phant. Semen freezing experiments were and ultimately allowing zoos not to conducted on ten ejaculate from one bull. permanent.” rely on forcing animals to breed. “If we Attempts to collect from the other Asian could use animals that were already elephant bulls resulted in samples that dead – even from 20 years ago – to were not suitable for freezing, or that could generate sperm and eggs then we can not be frozen at the time of collection1. use those individuals to create greater Te ten semen samples were evaluated and that with fresh semen in most cases. It is genetic diversity. I see it as being pos- then processed for freezing with various important to recognize the negative efects sible. I see no scientifc barrier,” Loring cryoprotectant to determine the best freez- of cryopreservation as it is still quite im- 3 says. ing technique suitable for Asian elephant perfect. Several of the cellular organelles of Despite some exceptions, many of semen. Te seven diferent concentra- sperm are enveloped by one or more mem- the frozen cells of certain species were tions of cryoprotectant that were used branes and it is known that membranes are not suitable for the long-term preser- were: ethylene glycol, propylene glycol, particularly vulnerable to survival during vation of undamaged DNA. Although trehalose, egg yolk, glycerol, and cryopro- cryopreservation. Sperm membranes certain species produce unsuccessful tectant-glycerol and Me2SO. In the study, afected by cryopreservation include the cells for cryopreservation, there are the scientists concluded that glycerol was plasma membrane, the outer acrosomal some species that produce ideal cells the best cryoprotectant for freezing sperm membrane and and mitochondrial mem- and work fantastically well. Scientists cells. According to the study, using glycerol branes.4 Tere are at least two diferent have successfully conducted cryopres- “increases the intracellular osmolarity and phases the sperm cell experiences during ervation on Asian elephants. Te Asian by that decrease cellular dehydration and freezing and thawing. Te frst relates to 1 elephant (Elephas maximus) worldwide shrinkage”. the efects of changing temperature and population is estimated to be at around Still, even with the best cryopreser- the second arises because of the formation 50,000-70,000 elephants, of which vation techniques, the post-thaw survival and dissolution of ice.4 Te sperm expe- approximately 15,000 are in captivity. rates for sperm cells are about 50%. As riences a cold shock, which traditionally Unfortunately, its population in the a consequence, fertility from artifcial refers to the extreme sensitivity to sudden wild is not reproducing at a sufcient insemination (AI) is more worse than cooling exhibited by spermatozoa. Te cell membrane becomes fragile in the cold and the chances of survival decreases. Second, ice crystals form in the cells that can cause the membranes to rupture when thawing. Te severity of the efects vary among species but are all dependent on the rate of cooling. Tus, cryopreservation is not entirely reliable for unnatural animal reproduction due to membrane vulnerability. In addition, not only is the cell’s membrane extremely vulnerable during freezing but there are also the possibility of contamination. Te DNA cells are kept frozen in a liquid nitrogen storage. Liquid nitrogen can also act as a carrier for viruses, bacteria, and fungi.2 If the liquid nitrogen is contaminated, then the cells will mostly be contaminated as well. Cryopreservation appears to be success- ful in some instances. However, in order to bring back extinct species, scientists need to fgure out the best method that Frozen DNA sample being extracted from storage

FALL 2016 | Berkeley Scientifc Journal 51 nary College, 1995.

IMAGE SOURCES

[1] Google images | https://lh3.googleus- ercontent.com/ePr0TgnTWEJV1F40Cx- rWXwHB63hsoPZ2B8_BYHAhtEuBg- Peww_f8iz0eatIADc-FdxE6Vw=s150 [2] Google Images | http://www.mmegi. bw/2016/june/27/phpE6qNCc.jpg [3] Google Images | https:// www.google.com/url?sa=i&rct=- j&q=&esrc=s&source=images&c- d=&ved=&url=http%3A%2F%2Fwww. trbimg.com%2Fimg-57e47b34%2Ftur- bine%2Fsdut-frozen-zoo-20160922%2F300%2F300x- 169&psig=AFQjCNHw9NULdtiTWv4oW KuFFf-TefT2_A&ust=1480312942147819

Scientist holding a frozen DNA sample works for all species so that we don’t waste in the frozen zoo at San Diego’s Insti- precious DNA when sperm cells are lost. tute for Consveration Research. Unfortunately, cryopreservation has not been proven to have long-lasting efects on all mammals, although fsh, reptiles, and many other species hatched through eggs have been farmed and have been successfully reintroduced into the wild. None- theless, cryopreservation appears to open many doors for the future of de-extinction and the survival of endangered species. However, there is still much to learn about cryopreservation before we can move on to the possibility of resurrecting extinct species.

REFERENCES

[1] Behr, Brita., Hermes, Robert., Hildeb- randt, Tomas B., Knieriem, Andreas., Kruse, Jürgen and Saragusty, Joseph. Suc- cessful cryopreservation of Asian elephant (Elephas maximus) spermatozoa. Animal Reproduction Science, 2008. [2] Burder, David W. Issues in Contami- nation and Temperature Variation in the Cryopreservation of Animal Cells and Tissues. BT&C, Inc. [3] Schultz, David. Should we bring extinct species back from the dead? Science Mag- azine, 2016. [4] Watson, P.F., Recent Developments and Concepts in the Cryopreservation of Spermatozoa and the Assessment of their Post-thawing Function. Department of Veterinary Basic Sciences, Royal Veteri-

52 Berkeley Scientifc Journal | FALL 2016