Improvement of bone marrow mononuclear cells cryopreservation methods to increase the efficiency of dendritic cell production.

ABSTRACT

Cryopreservation is now considered an integral part of the biotechnological process, exploiting different types of cells and tissues in clinical practice. Among them, dendritic cells (DCs) deserve special attention, notably the immature tolerogenic cells (tolDCs), which provide natural tolerance in humans and animals. High cryolability of tolDCs has necessitated the search for the methods that would provide cryopreservation of their precursors; those more resistant to negative effects of cryopreservation factors, in particular, bone marrow or peripheral blood mononuclear cells (MNCs). Based on this, the aim of our research was to optimize the cryopreservation conditions for mice bone marrow MNCs with further assessment of their ability to form tolDCs ex vivo. A cryopreservation mode for bone marrow MNCs has been developed which provides structural and functional completeness of tolDCs obtained from them ex vivo. The ability of DCs derived from cryopreserved MNCs by the developed mode to induce T-regulatory (FOXP3+) cells in vitro when co-cultured with CD4+-lymphocytes was shown.Tolerogenic properties of the DCs derived from cryopreserved MNCs are implemented by increasing the content of hsp70 heat shock proteins and the expression rate of glucocorticoid-induced leucine zipper (GILZ). DCs with increased tolerogenic activities, obtained by the developed cryopreservation regimen, can be used in treatment of autoimmune diseases. In this research we not only evaluated the qualitative characteristics and tolerogenic activity of DCs produced in vitro from cryopreserved MNCs, but also outlined the prospects of accumulating their reserves in low-temperature banks for clinical use.