By Sven Bulterijs for LongeCity, February 19, 2017

1. How has the cryopreservation procedure evolved since the first human was placed in cryostasis?

The most important element in the progress of cryopreservation procedures in cryonics is the progressive elimination of ice formation. When cryonics started, patients were often cryopreserved without any cryoprotection or very low concentrations of cryoprotectant. In the 1980’s and 1990’s organizations such as Alcor started adapting mainstream perfusion technologies to introduce high concentrations of cryoprotectants (such as glycerol) to mitigate ice formation. In 2000 Alcor formally introduced vitrification with the aim of eliminating freezing altogether.

2. Have changes in the procedure over the last decades (composition of cryoprotectant, rate of cooling, etc.) lead to a measurable decrease in the damage that occurs during vitrification?

Yes. The elimination of ice formation, which can be achieved in good cases, removes one major form of mechanical damage in the cryopreserved brain. One very attractive feature of a low-toxicity vitrification agent like M22 is that it does not require rapid cooling to prevent ice formation. Under good circumstances (no prior ischemia) it can also be used in whole-body patients without edema – a problem that seemed to plague prior DMSO-based cryoprotectants in cryonics. Elimination of ice formation and reduced toxicity has substantially reduced the degree of damage associated with cryopreservation.

3. Which foreseeable advances in the field of cryobiology do you believe will lead to improvements in how humans are preserved and decrease the damage done on the bodies?  

I foresee further advances in two areas; a more detailed understanding of the nature of cryoprotectant toxicity and the design of brain-optimized cryoprotectants. Cryoprotectant toxicity is currently the most formidable obstacle preventing reversible cryopreservation of complex mammalian organs. With the exception of the work of Dr. Greg Fahy and his colleagues at 21^st Century Medicine, it is rather surprising how little theoretical and experimental research has been done to illuminate the mechanisms of cryoprotectant toxicity. It is also increasingly recognized that the poor penetration of cryoprotectants across the blood-brain barrier causes dehydration of the brain. We need to develop brain-optimized vitrification solutions and/or identify better methods to deliver cryoprotectants to the brain without such significant changes in brain volume. Resolving these two issues will bring us much closer to reversible brain cryopreservation.

4. People have experimented in the past with a wide variety of antioxidants, chelators and membrane stabilizing molecules to reduce the damage to the body at the start of the procedure (so just after legal pronunciation of death). Have any of these been successful and are people still trying to find such substances to reduce damage at the early moments of the procedure?

I think it is important to recognize that all these anti-ischemia interventions are more important when there is a delay between pronouncement of death and the start of cryonics procedures. If there is a rapid and smooth transition between the two, immediate restoration of circulation, rapid induction of hypothermia, and aggressive anti-thrombotic therapy should be sufficient to maintain cerebral viability of the brain by contemporary medical criteria.

Our lab, Advanced Neural Biosciences, has collaborated with Alcor to conduct a rather comprehensive study of the effects of Alcor’s stabilization medications protocol and the most robust finding in this research has been that the combination of heparin and citrate allows for ice-free cryopreservation of the brain when these compounds are administered immediately after pronouncement of legal death. When medication administration is delayed by more than 15 minutes, things get more challenging and breakdown of the blood brain barrier and whole-body edema during cryoprotective perfusion is a typical outcome. Preventing edema of the patient during cryoprotective perfusion after prolonged periods of ischemia remains one of the most difficult research challenges to solve.

5. Preventing damage to the brain during cryopreservation is most definitely the top priority. However some damage remains inevitable. What evidence is there that the brain is not damaged by the cryopreservation process to such an extent that the information in it may be lost forever?

I can answer this in two ways. To start with, if we can eliminate ice formation in the brain, the damage associated with cryoprotectant toxicity is assumed to be mostly of a biochemical nature (i.e. denatured proteins) and does not alter the ultrastructure of the brain in a way that precludes inferring the original state. Cryoprotectant-induced dehydration of the brain is a little more of a wild card because we do not have much detailed information about the kind of ultrastructural changes associated with it. Hence, the priority to avoid the brain shrinking that is routinely observed in “good” cases. Ultimately, our incomplete knowledge of the neuroanatomical basis of identity, and about the exact capabilities and limits of future medicine, prompt us to be agnostic about the degree of damage that is still compatible with meaningful revival. Advocates of cryonics are sometimes accused of being too optimistic about future science, but perhaps skeptics are too pessimistic.

6. Do any changes take place in the bodies during cryogenic storage? And if such changes take place does that mean that the chance on successful reanimation will decrease over time?

No. To our knowledge (which is based on cryobiological studies and theoretical calculations), deterioration of patients stored at cryogenic temperatures should be non-existent or negligible. Things get a little bit more complicated when we store patients at intermediate temperatures (intermediate temperature storage or “ITS”) instead of liquid nitrogen temperatures. It has been suggested that nucleation may still occur slightly below the temperature where the vitrification solution turns into a glass (-123 degrees Celsius). At that temperature, however, nucleation does not translate into ice formation but it might create more challenging repair and revival scenarios.

7. Do you have any hypotheses on how the cryoprotectant could be removed from the body during the reanimation procedure and how hypoxic injury during this removal procedure could be prevented?

Roughly speaking, there are two distinct approaches to the repair and revival of cryonics patients. In the vision of researchers such as Robert Freitas and Ralph Merkle, a mature form of mechanical nanotechnology will be used to conduct the initial stages of repair and cryoprotectant removal at cryogenic temperatures. If this vision of nanotechnology is plausible, cryoprotectant can be removed while providing (local) metabolic and structural support to prevent damage or freezing. An alternative vision of nanomedicine will involve the use of biological repair machines such as modified viruses or modified white blood cells that operate using conventional diffusion-driven chemistry rather than molecular mechanical nanotechnology. Repair is more challenging in this biological scenario because tissue first needs to be warmed to temperatures at which the cryoprotectant solution inside cells and tissue becomes liquid.  This risks movement of damaged structures, possible growth of ice, and cryoprotectant toxicity accumulation occurring at the same time as repairs are being made.  To my knowledge, there have not been many serious studies of how such devices can operate and navigate through these problems at the same time.

8. What is in your opinion the chance that a cryopreserved person would be revived in a human state versus an uploaded version as uploading may be a way around irreparable cryopreservation damage?

I am personally partial to the idea of doing molecular level repairs through mechanical or biological nanomedicine because it does not require a paradigm shift in how we think about the nature of identity and consciousness. The feasibility of mind uploading is ultimately about the feasibility of substrate-independent minds and I do not think that the debates surrounding this can be resolved prior to empirical verification. In my opinion, the proposal of cryonics is intrinsically linked to the idea that the non-damaged state of the brain can be inferred from the damaged state through some form of molecular medicine. Many people feel quite comfortable with reconstruction of ancient DNA or forensic inference, but when it comes to cryonics, people tend to treat the brain in a somewhat superstitious fashion and cannot imagine forms of medicine that operate with molecular precision.

9. Even if reanimation after cryopreservation becomes technologically possible, what would make you believe that future generations will spend the money and resources on reanimating all people from cryostasis rather than just one or a few as an experiment?

This is an easier question to answer because it is the aim of cryonics organizations themselves to resuscitate their patients, not the general public, or curious scientists. The Alcor Life Extension Foundation parks a rather substantial portion of the cryopreservation fees in a so-called Patient Care Trust that should permit patients to be maintained in perpetuity (in theory) and revived when the technologies are available and affordable. Of course, if the technologies to revive cryonics patients will come to fruition, it seems quite reasonable to assume that the legal status of cryonics patients will also change and patients at cryonics organizations will be considered living people in a critical condition.

10. Do you see a mutual exchange of techniques and knowledge between the human cryopreservation field and the field of storing human biological samples (e.g. sperm, fertilized eggs, etc.)?

Yes. As a general rule, the obstacles that are faced by researchers of storage of biological samples and complex mammalian organs are the same obstacles that need to be overcome for reversible cryopreservation of humans (medical biostasis) as well. Any insights into the mechanisms of cryoprotectant toxicity, chilling injury, and the effects of cryopreservation on gene expression are of great relevance to cryonics. I should add, however, that I expect this exchange to be mutually beneficial. One of the least recognized and appreciated aspects about the field of cryonics is that researchers sympathetic to the idea of human cryopreservation have made meaningful and innovative contributions to mainstream fields such as cryobiology and cerebral resuscitation.

11. Cryogenic storage of genetic mutants in laboratory animals could reduce the cost of biomedical research. This is already a common procedure in the roundworm elegans. Are you aware of any research taking place that tries to expand cryogenic storage to other model organisms?

Natasha Vita-More, who conducted recent studies on the effects of vitrification on memory in C. elegans, has suggested that the next step would be a slightly more complex organism such as the Greenland Woolly Bear Caterpillar or the ozobranchid leech. One of the most common suggestions I get is to attempt suspended animation on a mouse or rat. This would definitely provide powerful proof of principle for the feasibility of human suspended animation, but I do not think that the challenges in achieving reversible biostasis in a small mammal are that much smaller than in humans. We would need to overcome the same obstacles: minimizing cryoprotectant toxicity, chilling injury, dehydration of the brain, ischemia during cooling, and cryoprotective perfusion, etc. The majority opinion in cryonics is to solve these individual problems more thoroughly before attempting reversible cryopreservation of a complete animal.

12. In his ruling on the 14-year old girl wanting cryonics, justice Peter Jackson stated that there’s a lack of regulation concerning cryonics. If the government would ask your advice on creating such regulations then what would you tell them?

I think the first thing I would recommend is that experts (which should include researchers and practitioners of the field) create a protocol to conduct cryonics as a hospital-based, elective, medical procedure. Reviewing the technical requirements and supporting evidence for cryonics will lead to a greater recognition of the need of improved legal protection for cryonics patients. Too often, cryonics is dismissed because people do not understand the conceptual arguments in favor of it, or its multi-disciplinary nature. In particular, the idea of molecular medicine is usually ignored in discussions about the (potential) damage of cryonics procedures. If regulations and protocols are created based on a dispassionate examination of the arguments and evidence in favor of cryonics, I think we do not necessarily need to fear regulation of the field.