Production of immunoregulatory molecules by induced erythroblasts at various stages of cell differentiation

   Introduction. Bone marrow erythroblasts produce a wide range of cytokines with opposite biological effects. This may be due to a change in the spectrum of production of immunoregulatory mediators during differentiation and small qualitative and quantitative differences in the spectrum of cytokines produced at each stage of differentiation, which may be important for the regulation of hemo- and immunopoiesis.   The aim. To study the spectrum of production of mediators by erythroblasts at different stages of differentiation.   Methods. Erythroblasts were obtained from CD34+ bone marrow cells of healthy donors in the presence of recombinant cytokines. Phenotype assessment was performed using flow cytometry for erythroid (CD45, CD71, CD235a, CD44) and lymphoid markers (CD3, CD4, CD8, CD16, CD19). Blocking of erythroblast differentiation at different stages was carried out using specific blocking monoclonal antibodies to melanocortin receptors (MCR) of types 1, 2 and 5. Cytokine analysis in conditioned erythroblast media was performed using the Bio-Plex Pro Human Cytokine 48-Plex Screening Panel (Bio-Rad Laboratories, USA). Cytokine production was analyzed using the CytokineExplore online tool.   Results. The resulting erythroblasts are divided into positive and negative populations according to the CD45 marker, carry markers of erythroid cells CD71, CD235a and do not express linear markers of lymphoid cells. In type 1 MCR blockage, polychromatophilic erythroblasts predominate, in type 2 MCR blockage, basophilic erythroblasts predominate, and in type 5 MCR blockage, orthochromatophilic erythroblasts accumulate. According to the production of cytokines, it was shown that when using any of the blocking antibodies, we obtain cells that differ qualitatively and quantitatively in a number of mediators from the initial population of induced erythroblasts.   Conclusion. Thus, we have shown qualitative and quantitative differences in the production of mediators by erythroblasts depending on the stage of differentiation, which can lead to different regulatory effects.