X-Message-Number: 33098
Date: Sat, 04 Dec 2010 16:52:58 -0500
From: 
Subject: Re: could argon/xenon improve vitrification solutions?

   Sorry if I am always raining on your parade, Doug, but
when I see misconceptions in your postings I feel the need
to set the record straight if there is to be hope of progress.
I do greatly appreciate the interest you show in the
possibility of reversible cryopreservation and the research
that you do toward this end.

http://www.cryonet.org/cgi-bin/dsp.cgi?msg=33097

All of the studies you cite except the xenon clathrate biostasis
paper deal with the subject of reducing ischemic damage by
substituting noble gas for oxygen. Despite the fact that a couple
of these papers involve hypothermia, this has no relevance
to improving vitrification solutions.

    I also doubt that xenon clathrate cryostasis is of any benefit
to vitrification, although it could be of value in cryopreservation
methods classically associated with freezing. The debate still rages
within the cryobiology community as to whether freezing or vitrification
is the best method of cryopreservation. Many single-cell and small
tissue specimens can be successfully cryopreserved in liquid nitrogen
using small amounts of cryoprotectant that reduce the ice formation
that occurs outside of cells. The small amounts of ice formed
can be tolerated in small tissue samples and cell suspensions, and
using only small amounts of cryoprotectant does not involve much
toxicity. Until less toxic cryoprotectants can be found, this is
the best approach for certain cell types. For more detail on
classical cryogenic cryopreservation using freezing, see

http://www.benbest.com/cryonics/cooling.html#classical

  These methods have no hope of ever being applicable to large
organs. Those ignorant of cryobiology often suggest that cells
burst upon freezing because ice has 9% greater volume than
water. That claim is false, because water first freezes
extracellularly, but it is nonetheless true that a 9% greater
volume of ice in the extracellular space will nonetheless
mechanically crush cells. Clathrate hydrates occupy a larger
volume than ice, so any attempt to form these clathrates in
an organ would result in even greater mechanical crushing.
As I said in my analysis of clathrate hydrates: "The volume of
a clathrate is greater than that of hexagonal ice containing
the same number of molecules ? even excluding the guest molecule
... clathrate formation in biological tissues is not an
alternative to vitrification, and would actually be expected
to cause more damage than freezing."

http://www.benbest.com/cryonics/viable.html#clathrates

Freezing techniques cannot complement vitrification, and
neither can clathrate formation. These techniques are
mutually exclusive. I note that in the xenon clathrate
cryostasis paper you cite

http://www.ncbi.nlm.nih.gov/pubmed/18787624

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2480575/

that 100 psi (6.8 atmospheres) pressure is used. High
pressure methods can be used to reduce ice formation
due to the 9% greater volume of ice over water, but in
this case high pressure is probably mostly to induce
the xenon to form the clathrate. It was once necessary
to use 1,000 atmospheres of pressure with VS4
vitrification solution (higher pressures were too
damaging), until the development of VS41A
(VS4 for 1 atmosphere) vitrification solution.

  The bottom line is that although xenon clathrates may
be useful for cryogenic temperature cryopreservation of
cell suspensions and small tissue samples, it cannot
be applied to cryogenic organ cryopreservation or
cryonics. Reduction of cryoprotectant toxicity
remains the number one priority. There are methods
that complement vitrification, such as ice blockers
and high pressures, but clathrate formation is not
one of these methods.

       -- Ben Best

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