Cryogenic Burials & Preservation

1. Cryogenic Burials & Preservation Maxim Gorshkov Visit my website for this slideshow: www.mgorshkov.com/mcgill/biol112

2. Cryonics • The low temperature preservation of humans and animals • Goal: Successfully preserve human being, revive or take out of preservation at later point. • Fairly theoretical concept: long-term memory, personality, and identity stored in cell structures that do not require continuous brain activity to survive.

3. Three Principles of Cryonics • Life can be stopped and restarted if the structure is properly preserved • Certain organs (ex. Heart) can be cooled until stopped, restarted within the hour. • Structures can be preserved well by Vitrification • Rather than freezing, certain technique can be performed • Methods for repairing structures are now foreseen. • Nanotechnology and the like now imminent and relevant to Cryopreservation

4. Cryopreservation • Basic testing involving Cyanobacteria • Cells in Cyanobacterial suffer stress osmotic stress/ice crystal damage during thawing/freezing processes • Extrapolation to all life: damage with all living cells • Reduce effects with Cryoprotective compounds • Specifically Glycerol or ~5% Methanol for Cyanobacteria

5. Cryopreservation: Applicability to Animals/Humans • Cyanobacteria represents most living cells within animals • Researchers began in the late 1990s to work with composition tissues such as cartilage. • Pioneered further Cryopreservation research

6. Cryopreservation: Composite Tissue • Rats were anesthetised and two flaps were marked on the epigastric region. • Incision was made around one of the flaps • Tissue removed was stored at 4 degrees until preserved

7. Cryopreservation: Composite Tissue (2) • To study effects: • Skin defect was formed, preserved flap was interposed and sutured • Conclusions: • Potential expansion of technique but only 2/17 cryopreserved rats survived unique experiment. • Key factors were investigated to include the preservation agent and the warming rate.

8. Vitrification • Most popular/researched technique for cryopreservation • Based on premise that amphibians/insects tolerate varying amount of freezing • Since the viscosity of water is low, it can be vitrified by extremely rapid “flash-freezing” • The new structure in the polymer-like form is the “vitrified form” rather than the crystalline structure of regular frozen water.

9. Vitrification: Cryoprotectors • Theory: cryoprotectors would work as well as flash-freezing (or at least aid) • Study done in 1949 in which glycerol was introduced to protect bull sperm against freezing, further proven with red blood cells later on • DMSO also introduced as cryoprotectant, useful and easily passes through membranes • Mouse embryos preserved though combination of DMSO and glycerol

10. Vitrification: A Simulation Freezing Without Cryoprotector

11. Vitrification: A Simulation Freezing With Cryoprotector

12. Cryogenics Today • We’ve seen that freezing/warming rates of a sample affect the viability of the sample • Research has been done at University of Tennesse and has achieved a reasonable survival rate of mouse oocytes. • Through the vitrification cooling to -196 degrees Celsius and then warming to room temperature a sophisticated experiment was performed

13. Survival Rate of Mouse Oocytes • Cooling/warming rates varied to investigate survival rate • Procedure: • Mature mice made to superovulate and the oocytees were collected • Oocytes were vetrified in a solution of Ethylene Glyocol, Acetamide, and Ficoll. • Oocytes were collected when warm and the viability was assessed.

14. Survival Rate of Mouse Oocytes (2) • Based on the results, 70-80% of sample surivied regardless of cooling rate. • Slowest warming rate had loest percentage of sample survival.

17. Survival Rate of Mouse Oocytes: Conclusions • High sensitivity of suvival to warming rates  crystallization of intracellular structues during warming or growth responsible for lethality • Much more potential available for growth in the field based on this study and others.

18. Why is it a Burial? • In the United States, cryonics can only be legally performed on humans after they have been pronounced legally dead as otherwise it would count as murder or assisted suicide. • Ideally this is performed with minutes of a cardiac arrest

19. Ethical/Social Considerations • Given that a company in US offers Cryopreservation for $80,000/$200,000… • Selfish/irresponsible use of money which can provide a potential equivalent amount of money to efforts in developing countries • Social implications for Cryopreserved person, be that they are preserved: culture shock, alienation. • Big leap forward in science and can also open up many new directions with implications in many fields.

20. Given the choice, would you want to be Cryopreserved?

21. References • Mazur P, Seki S. Survival of mouse oocytes after being cooled in a vitrification solution to −196 °C at 95° to 70,000 °C/min and warmed at 610° to 118,000 °C/min: A new paradigm for cryopreservation by vitrification. Cryobiology. 2011;62(1);1-7. • Jacobsen IA, Pegg DE, et al. Cryopreservation of organs: A review. Cryobiology. 1984;21(5):377-384. • Cryopreservation of Cyanobacteria [homepage on the Internet]. Austin: University of Austin; [cited 2011 Feb 28]. Available from: http://www-cyanosite.bio.purdue.edu/protocols/cryo.html. • Cui X, Gao D, Vasconez H, Rinker B. Cryopreservation of composite tissues and transplantation: Preliminary studies. Cryobiology. 2007;55(3);295-304. • Merkle RC. The technical feasibility of cryonics. Medical Hypotheses. 1992;39(1);6-16 • Cryonics: Alcor Life Extension Foundation [homepage on the Internet]. United States of America: Alcor Life Extension Foundation. Available from http://www.alcor.org. • Vitrification in Cryonics [homepage on the Internet]. Available from http://www.benbest.com/cryonics/vitrify.html.