[Immunohistochemical method of megakaryocytic lineage staining in bone marrow biopsy specimens as an additional pathomorphological differential diagnostic sign of primary myelofibrosis and essential thrombocythemia].

Abstract

Objective: To evaluate and compare morphometric and histotopographic characteristics of megakaryocytic lineage in preparations stained with H&E or antibodies to CD42b in diagnostic trepanobioptates of bone marrow of patients with primary myelofibrosis and essential thrombocythemia with JAK2 or CALR mutation. Analyze the dimensions and quantity of CD42b-positive megakaryocytes in 1 mm² area of section and assess suitability of these parameters as an additional differential pathomorphological criterion.

Material and methods: 108 trephine biopsies of the bone marrow from patients with primary myelofibrosis (N=53) and essential thrombocythemia (N=55) with JAK2 or CALR mutation were selected. Digitized bone marrow slides stained with H&E or antibodies to CD42b (clone EP409) were the object of study. In every sample the average values of perimeter and area of megakaryocytes were analyzed, as well as the average number of megakaryocytes in 1 mm² area of myeloid tissue section. Logistic regression analysis was used to describe the relationship between CD42b-positive megakaryocyte characteristics and disease (primary myelofibrosis or essential thrombocythemia).

Results: Immunohistochemical examination of bone marrow biopsy specimens using antibodies to CD42b in comparison with H&E staining allows to multiply the number of identifiable megakaryocytes in myeloid tissue by 3.5-4 times (p<0.0001). Statistically significant differences in the mean values of the number of megakaryocytes in 1 mm² of the section area and megakaryocyte perimeter in patients with primary myelofibrosis and essential thrombocythemia have been demonstrated. ROC analysis (AUC=0.84, 95% CI 0.7782-0.9199) justifies the inclusion of the average perimeter size of CD42b-positive megakaryocytes and their number in 1 mm² of the section area in the differential diagnostic panel as an additional pathomorphological criterion.

Conclusion: The revealed statistically significant differences in quantitative and geometric characteristics of megakaryocytes allowed to calculate differential threshold values of characteristics of megakaryocytic lineage of myeloid tissue in diagnostic trepanobioptates of bone marrow from patients with primary myelofibrosis and essential thrombocythemia. Counting the number of CD42b-positive megakaryocytes in one field of view at a magnification of 400 times was proposed as an additional pathomorphological differential-diagnostic sign.