Immunohistochemical study of Ulex europaeus agglutinin 1 (UEA-1) binding of megakaryocytes in bone marrow biopsy specimens: demonstration of heterogeneity in staining pattern reflecting the stages of differentiation.

During differentiation, megakaryocytes undergo nuclear endoreplication, an increase in cell size, cytoplasmic granulation, and release of platelets. The changes in highly lobulated nuclei with varying degree of polyploidy and increasing cell size are easily recognized morphologically. However, the actual cytoplasmic changes are more difficult to perceive morphologically. With the peroxidase-antiperoxidase (PAP) method using UEA-1 as the binding protein to the alpha-L-fucose of glycoprotein synthesized by megakaryocytes, we observed significant variation in cytoplasmic staining of megakaryocytes in routinely processed bone marrow biopsy sections. A total of 3344 megakaryocytes in bone marrow sections from 10 patients with nonhematologic diseases and from 10 patients with idiopathic thrombocytopenic purpura (ITP) was studied. According to the intensity and pattern of cytoplasmic staining, we divided megakaryocytes into at least six groups: (1) low granular (LG), (2) diffuse granular (DG), (3) diffuse dense granular (DDG), (4) marginal granular (MG), (5) denuded (DMK), and (6) endomitotic (EndoM). Most of the megakaryocytes were DG (mean, 42.75% +/- 19.21%) and DDG (mean, 50.25% +/- 21.23%). In correlation with nuclear morphology and cell size, it appears that substances binding to UEA-1 are located in the paranuclear region in early megakaryocytes and produce a low granular focal staining pattern (LG cells). Next, the granules spread throughout the cytoplasm (DG cells) and increase in quantity (DDG). This is followed by migration of granules to the periphery of the cytoplasm (MG cells) and is associated with the liberation of platelets and eventual formation of DMK megakaryocytes. Endomitosis, regulated by unknown factors, occurred in the MG stage. In comparing the group with nonhematologic disease (mean DG, 35.4% +/- 18.48%; DDG, 58.4% +/- 21.8%) and the group with ITP (mean DG, 50.1% +/- 17.82%; DDG, 42.1% +/- 18.12%), we found an increasing proportion of DG megakaryocytes in ITP, which suggests a left-shifted maturation of megakaryocytes. By understanding the staining pattern seen in the different stages of megakaryocytic differentiation, UEA-1 staining may be a practical method for studying megakaryocytopoiesis in routinely processed paraffin sections of bone marrow biopsy samples.