1Institute of Cell Biophysics, RAS, Pushchino, Russia;
2Institute of Theoretical and Experimental Biophysics RAS, Pushchino, Russia;
3Shemyakin - Ovchinnikov Institute of Bioorganic Chemistry RAS, Moscow, Russia; email@example.com
The presence of neuroprotective factors in the cryoprotective mixtures added on the various stages during cryopreservation of nerve cells and tissues is very important for protection from extra low temperatures and for the successful recovery of the biological material after long-term preservation in cryobanks. The processes of recrystallization during process of thawing affect on the shape and size of the micro-particles in cryoprotective solutions. This in turn leads to cryodamage of neuronal cells and tissues.
The aim of this study was to reveal the possible neuroprotective effect of the peptide Thr-Ser-Lys-Tyr (TSKY) during process of the snail brain thawing from extra low temperatures. This peptide was isolated from brain of hibernating ground squirrels (Spermophillus undulatus).
Freezing – thawing of the isolated snail brain of Lymnea stagnalis was carried on with the rate of 400-500°С/min in presence of 2 M DMSO ( Gakhova et al.,1989). The peptide TSKY was added only in the solution for thawing and washing out of ganglia from cryoprotectants. The physiological activity of the isolated neurons after thawing of the brain was estimated by counting of number of the regenerated neuritis in neurons.
The results indicated that total number of living neurons in cell culture (estimated by trypan blue), which was isolated from thawing brains, was increased by 2 times as compared with non- treated brains (p < 0.0004). However, the number of viable neurons with neuritis was decreased by 10% (p < 0.033). We conclude that the peptide TSKY could protect neurons from negative effect of ice microparticles by decreasing the sensitivity neurons to cryodamages.
This study was supported by grant from the RFBR (№10-04-01319-а).