Studies On The Cryopreservation Of Embryonic Cerebral Cells By Vitrification And The Effect To The Vitality Of Cells

In the present study, pregnant SD rats (17~19 days) were used for the studies on the embryonic cerebral cells (ECC), which were cryopreserved by the vitrification method. The characteristic of vitrification solutions were analyzed by the toxicity of cryoprotectants on ECC and the process of pre-equilibration. The suitable vitrification solution and procedure were selected due to the recovery rates, the survival rates , the vitality of SOD and SDH in mitochondria and the structure of the cell membrane. And the factors affecting the viability of ECC were analyzed. The results of experiments showed that:(l)The analysis of the characteristic of vitrification solutions:The nonosmosiscryoprotectants (Ficoll and saccharose) had no toxicity effect to ECC. And the toxicity effects of cryoprotectants was from DMSO and EG, and the effect became more severe when the the time of pre-equilibration was prolonged and the concentration of the solutions were increased. And it also showed that the toxicity of DMSO is intenser than that of EG, however, DMSO had the excellent ability of vitrification .So the reasonable combination of DMSO and EG can achieve a better state of vitrification of ECC.(2) The selection of the vitrification solution: According to the experiment results, VS2 solution, consisted of 15%EG (v/v), 15%DMSO (w/v), 21 % Ficoll and 0.3 5M saccharose. After being cryopreserved for 3,7,15,30 and 60 days, the recovery rate was86.22 2.73%, 79.61 2.48%, 69.46 2.68%, 52.85 2.51% and51.29 3.19% respectively. The survival rates and the physiologic parametes after 30 days' cryopreservation were steady relatively.(3) The changes of the structures and the functions of the vitrified ECC: We have studied the vitality of SOD and SDH in mitochondria and the structure of the cell membrane and all the facts showed that in the process of cryopreservation the damage of the membrane and the depression of the viability of SDH were remarkable before 15 days of cryopreservation. the physiologic parametes after 30 days' cryopreservation were steady relatively. The results indicated that the cryopreservation by vitrification method was applicable to ECC for long-termpreservation.In the spite of the toxicity of cryoprotectants and the damage to ECC in theprocess of cryopreservation. The technique of vitrification still may maintain good vitality of ECC. It is indicated that cryopreservation of ECC by vitrification has somepractical value...