X-Message-Number: 31104
Date: Wed, 8 Oct 2008 09:40:51 -0700 (PDT)
From:
Subject: The Cryonics Institute Makes Another Technical Disclosure I
Message #31073 Date: Mon, 22 Sep 2008 00:34:33 -0400
From:
Subject: The Cryonics Institute Makes Another Technical Disclosure
>In February 2007 the Cryonics Institute disclosed the formula of
its vitrification solution CI-VM-1 developed by CI staff cryobiologist
Dr. Yuri Pichugin. Now in September 2008 CI (and the Immortalist
Society) is disclosing the Blood-Brain Barrier modifier which
Dr. Pichugin had spent much of 2007 perfecting and validating.
Details can be found on the Cryonics Institute website:
http://www.cryonics.org/research/BBB_Modifier_PPA.html
-- Ben Best
<
[Ben: Modifying the blood brain barrier would be essential to enable good
permeation of high molecular weight freeze protective substances. such as green
tea catechins, and kaempferol-7-O-glucoside. There exists the possibility that
good permeation of these extremely powerful but nonstandard cryoprotectants,
could enable a lowering of the molarity of vitrification solutions to the point
that cryoprotectant toxicity is no longer a significant factor.]
Cryobiology. 2008 Sep 15. [Epub ahead of print] Improved cryopreservation by
diluted vitrification solution with supercooling-facilitating flavonol
glycoside.
Kami D, Kasuga J, Arakawa K, Fujikawa S. National Agricultural Research
Center for Hokkaido Region, Sapporo 062-8555, Japan; Research Faculty and
Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589,
Japan.
The effect of kaempferol-7-O-glucoside (KF7G), one of the
supercooling-facilitating flavonol glycosides which was originally found in
deep supercooling xylem parenchyma cells of the katsura tree and was found
to exhibit the highest level of supercooling-facilitating activity among
reported substances, was examined for successful cryopreservation by
vitrification procedures, with the aim of determining the possibility of
using diluted vitrification solution (VS) to reduce cryoprotectant toxicity
and also to inhibit nucleation at practical cooling and rewarming by the
effect of supplemental KF7G. Examination was performed using shoot apices of
cranberry and plant vitrification solution 2 (PVS2) with dilution.
Vitrification procedures using the original concentration (100%) of PVS2
caused serious injury during treatment with PVS2 and resulted in no regrowth
after cooling and rewarming (cryopreservation). Dilution of the
concentration of PVS2 to 75% or 50% (with the same proportions of
constituents) significantly reduced injury by PVS2 treatment, but regrowth
was poor after cryopreservation. It is thought that dilution of PVS2 reduced
injury by cryoprotectant toxicity, but such dilution caused nucleation
during cooling and/or rewarming, resulting in poor survival. On the other
hand, addition of 0.5mg/ml (0.05% w/v) KF7G to the diluted PVS2 resulted in
significantly (p<0.05) higher regrowth rates after cryopreservation. It is
thought that addition of supercooling-facilitating KF7G induced
vitrification even in diluted PVS2 probably due to inhibition of ice
nucleation during cooling and rewarming and consequently resulted in higher
regrowth. The results of the present study indicate the possibility that
concentrations of routinely used VSs can be reduced by adding
supercooling-facilitating KF7G, by which more successful cryopreservation
might be achieved for a wide variety of biological materials.
PMID: 18824164
Plant Cell Environ. 2008 Sep;31(9):1335-48. Epub 2008 Jun 3.
Deep supercooling xylem parenchyma cells of katsura tree (Cercidiphyllum
japonicum) contain flavonol glycosides exhibiting high anti-ice nucleation
activity.
Kasuga J, Hashidoko Y, Nishioka A, Yoshiba M, Arakawa K, Fujikawa S.
Research Faculty and Graduate School of Agriculture, Hokkaido University,
Sapporo, Japan.
Xylem parenchyma cells (XPCs) of boreal hardwood species adapt to
sub-freezing temperatures by deep supercooling to maintain a liquid state of
intracellular water near -40 degrees C. Our previous study found that crude
xylem extracts from such tree species exhibited anti-ice nucleation
activity to promote supercooling of water. In the present study, thus, we
attempted to identify the causative substances of supercooling. Crude xylem
extracts from katsura tree (Cercidiphyllum japonicum), of which XPCs
exhibited deep supercooling to -40 degrees C, were prepared by methanol
extraction. The crude extracts were purified by liquid-liquid extraction and
then by silica gel column chromatography. Although all the fractions
obtained after each purification step exhibited some levels of anti-ice
nucleation activity, only the most active fraction was retained to proceed
to the subsequent level of purification. High-performance liquid
chromatography (HPLC) analysis of a fraction with the highest level of
activity revealed four peaks with high levels of anti-ice nucleation
activity in the range of 2.8-9.0 degrees C. Ultraviolet (UV), mass and
nuclear magnetic resonance (NMR) spectra revealed that these four peaks
corresponded to quercetin-3-O-beta-glucoside (Q3G),
kaempferol-7-O-beta-glucoside (K7G), 8-methoxykaempferol-3-O-beta-glucoside
(8MK3G) and kaempferol-3-O-beta-glucoside (K3G). Microscopic observations
confirmed the presence of flavonoids in cytoplasms of XPCs. These results
suggest that diverse kinds of anti-ice nucleation substances, including
flavonol glycosides, may have important roles in deep supercooling of XPCs.
PMID: 18518920
Transplantation. 2006 Jan 27;81(2):231-9.
Improved cold preservation of kidney tubular cells by means of adding
bioflavonoids to organ preservation solutions.
Ahlenstiel T, Burkhardt G, Kohler H, Kuhlmann MK. Department of Medicine,
Division of Nephrology and Hypertension, University Hospital of Saarland,
Homburg/Saar, Germany.
BACKGROUND: Cold ischemia and reperfusion during renal transplantation
result in release of reactive oxygen species. The aim of this study is to
examine whether cold storage induced cell injury can be ameliorated by
adding flavonoids directly to preservation solutions. METHODS: Cultured
renal tubular epithelial cells (LLC-PK1) were stored in University of
Wisconsin (UW) or Euro-Collins (EC) solution at 4 degrees C for 20 hours.
Preservation solutions were supplemented with various flavonoids. After
rewarming, structural and metabolic cell integrity was measured by lactate
dehydrogenase (LDH) release and MTT-test, and lipid peroxidation was
assessed from generation of thiobarbituric acid-reactive substances (TBARS).
RESULTS: Twenty hours of cold storage resulted in a substantial loss of
cell viability in both preservation solutions (in EC: LDH release
92.4+/-2.7%; MTT-test 0.5+/-0.7%). Addition of luteolin, quercetin,
kempferol, fisetin, myricetin, morin, catechin, and silibinin significantly
reduced cell injury (for luteolin in EC: LDH release 2.4+/-1.6%; MTT-test
110.3+/-10.4%, P<0.01; TBARS-production (related to cold stored control
cells) 8.9+/-2.6%). No cytoprotection was found for apigenin, naringenin,
and rutin. Protective potency of flavonoids depends on number of
hydroxyl-substituents and lipophilicity of the diphenylpyran compounds.
CONCLUSION: Cold storage induced injury of renal tubular cells was
substantially ameliorated by adding selected flavonoids directly to
preservation solutions.
PMID: 16436967
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