Kisielova H. Rationale for the use of tolerogenic dendritic cells obtained from cryopreserved progenitors in the treatment of autoimmune diseases (experimental study).

The dissertation is devoted to the study of the features of the cryopreservation effect under different regimes on the structural and functional characteristics of bone marrow mononuclear cells (MNC) and dendritic cells (DC) obtained from them in vitro. Considerable attention is paid to the assessment of the tolerogenic potential of DCs obtained from cryopreserved MNCs, namely, the ability to minimize the manifestations of adjuvant arthritis (AA) in mice as an experimental model of rheumatoid arthritis (RA) in humans. The following modern research methods were used in the work: cryopreservation, in vitro cultivation, flow cytofluorimetry, real-time PCR, immunomagnetic cell sorting, inverted and light microscopy. A method of isolating MNCs from the bone marrow of 20-week-old SBA/H mice in a trasograph gradient was developed, which allows obtaining more than 40% of cells, among which 12% were CD14+ monocytes - DC precursors. It was established that the differentiation of monocytes into immature DCs is accompanied by the appearance of a marker characteristic for these cells - CD11b, the loss of the CD14 antigen and the minimization of the level of expression of costimulatory molecules CD80, CD86. It has been demonstrated that an important component of the pathogenesis of AA is the dysregulation of the function of the immune system (IS) against the background of an imbalance of the cytokine profile with a predominant increase in the content of pro-inflammatory cytokines (IL-6, TNF-α, IFN-γ) and dysfunction of the T-regulatory (Treg) link of immunity. Such changes were accompanied by an increase in clinical and laboratory indicators: the level of sialic acids, seromucoids, and the index of arthritis (IA). The influence of different modes (P) of cryopreservation under the protection of 10% DMSO with a slow cooling rate was studied: 1 degree / min to - 80ºС (P1); up to - 40ºС (P2); to -25ºС (P3) followed by immersion in liquid nitrogen (-196ºС) in each case for the structural and functional properties of bone marrow MNC. It is shown that changing the conditions of cryopreservation can increase the tolerogenic potential of DCs obtained for immunotherapy.