X-Message-Number: 7822
Date: Fri, 7 Mar 1997 22:21:41 -0800 (PST)
From: Doug Skrecky <>
Subject: frozen fetal mouse hearts Part II

    Fetal mouse hearts were revived successfully from liquid nitrogen
 storage  back in 1974 by the MRC Transplantation Group at the University
 of Alberta in Edmonton, Canada. Cryopreservation fluid included 10% DMSO
 and 10% FCS (fetal calf serum) in Hepes buffer. Deletion of either of
 these two components eliminated survival. In all 59% of 54 treated hearts
 used showed strong electrical activity after being thawed out.

 Reference: Cryobiology 11: 28-32 1974

    After reading some of the comments I recieved about the above abstract
 on successful revival of frozen fetal mouse hearts I now feel my abstract
 was a little too brief and I would like to add some further details here.
 Here's why I think the solution to every cryonicists dream of reversible
 cryopreservation was to all intents and purposes solved over 30 years
 ago.
    The fetal mouse hearts used were about 1 mm in diameter. Now one
 problem extrapolating from small to large tissue samples is that large
 samples can not be cooled as rapidly as small ones. However high cooling
 rates were not employed so this is not a problem. The freezing rate was
 maintained by a regulated thermocouple between 0.5 and 0.7 C/min down to
 -100 C. Then the samples were exposed to liquid nitrogen vapour and
 further cooled to -196 C at 5 to 10 C/min. The hearts were stored at this
 temperature for 72 to 216 hours before being rewarmed.
    Since Biopreservation claims to possess the technology capable of
 cooling/heating a human body at up to 10 C/min, their technology should
 in principle be able to maintain good viability in a wide variety of
 tissues in entire human bodies down to liquid nitrogen temperatures.
 However there is a hook that I have to mention here.
    Although relatively slow rates of cooling were employed, the heating
 was much more rapid. Two heating techniques were used. A warm bath was
 used to heat the hearts at about 150 C/min and as an alternative
 microwaves were used to heat at about 200 C/min. How much slower the
 heating could have been without compromising viability is unknown here
 since it was not tested. There was no difference between 150 and 200
 C/min. I suspect, but can not prove that 10 C/min may be too slow with
 the solutions used. This was either A) Eagle's minimal essential medium
 containing Hepes buffer, 10% fetal calf serum and 10% DMSO, or B) McCoy's
 5a medium containing Hepes buffer, 10% fetal calf serum and 10% DMSO.
    Two other solutions failed to preserve heart function. These were C)
 Cross solution containing 10% DMSO, but no fetal calf serum or other
 proteins and D) the same as solution A, but without the DMSO. Based on
 the excellent results with solutions A & B and negative results with C &
 D it seems that both fetal calf serum proteins and DMSO were required for
 success.
    With possibly a little improvement (1) in the solution I strongly
 suspect that reversible whole body resuscitation from liquid nitrogen
 storage should be possible in the near future.

 (1) Ethylene glycol is less toxic to hearts than DMSO for example.


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