Improving of cryoprotectant media for human cord blood nucleated cells cryopreservation.

Recently during treatment of many diseases the cryopreserved suspension of cord blood (CB) nucleated cells (NCs, CD45+) has been used as a preparation containing hematopoietic progenitor / stem cells (HPCs, CD34+). Efficiency of their application is mainly determined by the preservation of NCs structural and functional integrity after thawing. In this context, the purpose of the study consisted in comprehensive assessment of the structural and functional states of various populations of human CB NCs including HPCs before and after cryopreservation. Nucleated cells isolation in dextran solution and subsequent freezing with 5 % dimethyl sulfoxide (DMSO) allows to keep viable more than 80 % CD45+- and 90 % CD34+-cells. Among all CD45+-cells lymphocytes and monocytes are the most cryoresistant: more than 95 % and 90 % viable cells correspondingly. Intracellular content of reactive oxygen species (ROS) increased in average by 19 % (DCF-labeled CD45+-cells) after cryopreservation. Prevention of the ROS production by adding of 10-15 mM N-acetyl-L-cysteine (NAC) into cryoprotective medium reduced the number of DCF-labeled CB NCs and improved preservation and viability level. Thus, NAC can promote antioxidant processes and prevent oxidative stress in cells upon cryopreservation that improves their safety and viability.