Immunophenotypic study of bone marrow erythrokaryocyte differentiation in patients with diffuse large B-cell lymphoma

Introduction. Previously we»ve shown that there are typical changes in haematopoiesis in diffuse large B-cell lymphoma patients (DLBCL). Attention was paid to the elevation of oxyphylic erythroblasts, as this sign was of prognostic significance. The aim of this work was to see
if there is possible to identify bone marrow erythrokaryocyte maturation stages using flow cytometry method.

Materials and methods. Study was done in 60 DLBCL patients. Elevation of oxyphilic erythrobast levels was noted in 50 patients (83,3 %). Flow cytometric study of bone marrow were done in 30 patients. We used monoclonal antibodies to Glycophorin A, CD45, CD36, CD117, CD71, CD105, CD34, HLA-DR, as well as nuclear staining for SYTO 16, SYTO 41.

Results. Use of Glycophorin A together with of SYTO 16 or SYTO 41 is effective for identification of erythrocaryocytes, main proportion of which is represented by CD71+ and Glycophorin A+ cells, which does not express any precursor markers. In CD45-negative cellular elements there are as well CD71- glycophorin А– cells, which represents mainly precursors – CD34+ CD117+ HLA-DR+/–. Erythro caryocytes with low levels of glycophorin A expression and prominent expression of CD71 (committed to erythroid differentiation) show slight expresiion of CD117 and HLA-DR, but CD34 is completely absent. The informative method for characterization of most mature erythrocaryocyes was the approach based on identification of erythrocaryocytes according to CD36 expression on cels with low SSC (in that case usage of nuclear stain, CD45 and glycophorin A may be omitted as CD36 does not stain erythrocytes). After peak of CD36, CD71, CD105 expression levels of these antigens are declining. Namely that fraction represents most mature erythrocaryocytes. Cor relation of the percentages of that cells with oxyphilic normoblasts (identified morphologically) was near to significant value.