Clonal Expression of the Tn Antigen in Erythroid and Granulocyte Colonies and Its Application to Determination of the Clonality of the Human Megakaryocyte Colony Assay

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Abstract

To evaluate whether exposure of Tn determinants at the surface of human erythrocytes, platelets, and granulocytes could arise from a somatic mutation in a hemopoietic stem cell, burst-forming unit erythroid (BFU-E) colonies, colony-forming unit granulocyte-macrophage (CFU-GM), and colony-forming unit-eosinophil (CFU-Eo) were grown from a blood group O patient with a typical Tn syndrome displaying two distinct populations (Tn+ and Tn-) of platelets, granulocytes, and erythrocytes. A large number of colonies was observed. Individual colonies were studied with a fluorescent conjugate of Helix pomatia agglutinin (HPA). A sizeable fraction of each of the erythroid and granulocytic colonies appeared to consist exclusively of either HPA-positive or HPA-negative cells, thereby demonstrating the clonal origin of those exhibiting the Tn marker. Similar results were obtained from a second patient. These findings establish that the HPA labeling of Tn cells is an accurate marker permitting assessment of the clonality of the human megakaryocyte (MK) colony assay. For the study of MK cultures a double-staining procedure using the HPA lectin and a monoclonal antiplatelet antibody (J-15) was applied in situ to identify all MK constituting a colony. Our results, obtained in studies of 133 MK colonies, provide definitive evidence that the human MK colony assay is clonal because all MK colonies were exclusively composed of Tn+ and Tn- MK. Furthermore, the distribution of MK within a single colony was shown to be seminormal with a mean at 6 MK, isolated MK typically being absent in culture.

Authors

William Vainchenker, Ugo Testa, Jeanne Françoise Deschamps, Annie Henri, Monique Titeux, Janine Breton-Gorius, Henri Rochant, Douglas Lee, Jean-Pierre Cartron