Endogenous Megakaryocytic Colony Formation and Thrombopoietin Sensitivity of Megakaryocytic Progenitor Cells Are Useful to Distinguish Between Essential Thrombocythemia and Reactive Thrombocytosis

Abstract

Diagnosis of essential thrombocythemia (ET) is controversial and remains mainly an exclusion diagnosis. Endogenous megakaryocyte colony (EMC) formation have been largely evaluated to identify specific criteria for ET, but results are impeded by the lack of medium standardization. We evaluated megakaryocyte (MK) colony formation in a serum-free collagen-based medium, without cytokine and in the presence of various concentrations of thrombopoietin (TPO). Thirty-six bone marrows from patients diagnosed with ET (n = 11), polycythemia vera (PV; n = 12), reactive thrombocytosis (RT; n = 6) and healthy donors (n = 7) were assessed. We demonstrate that 11 out 11 of the ET patients had spontaneous megakaryocyte colony-forming unit (CFU-MK) formation, in contrast to none of the RT patients and healthy donors. MK progenitors from ET patients remained responsive to TPO, because exogenous addition of TPO significantly increased cloning efficiency. Moreover, at low doses of TPO (0.5 ng/ml and 5 ng/ml), the number of positive cultures and mean number of TPO stimulated CFU-MK were significantly higher in cultures of cells from patients with ET than in patients with RT. In summary, we have described a standardized serum-free, collagen-based assay that allows differential diagnosis of ET and RT, according to endogenous CFU-MK formation and sensitivity to TPO.

Spontaneous megakaryocytic colony formation does not discriminate between essential thrombocythemia and polycythemia vera

“…In contrast to our present data, various studies [3,5,6] have reported a lower percentage of positive CFU-MK assays in PV (ranging from 33 to 55%) compared to that occurring in ET. This may be linked to the variety of CFU-MK culture methods used, which is well known to be a source of difficulty for CFU-MK assay standardization [1].…”

“…Similarly, the percentage of endogenous CFU-MK growth and the absolute count of spontaneous and cytokine-dependent megakaryocytic colonies were not significantly different in ET and PV patients. As previously reported [3,4,6], we failed to detect endogenous CFU-MK growth in healthy individuals or in patients with reactive thrombocytosis (data not shown). Taken together, our data suggest clearly that the CFU-MK assay is useful to detect not only ET, but also PV, without discriminating between these two MPDs, however.…”

Endogenous megakaryocytic colonies underline association between megakaryocytes and calreticulin mutations in essential thrombocythemia

“…Linear regression was done according to type of CALR mutations (---for type 1, ▪▪▪ for type 2 and ----for "variants"). Julie Mondet, 1,2 Ji-Hye Park, 3 Audrey Menard, 4 Christophe Marzac, 5,6 Serge Carillo, 7,8 Emmanuel Pourcelot, 1,2 Francois Girodon, 3,9 Xenia Cabagnols, 10,11 Laurence Lodé, 4,12 Nuria Socoro, 1,2 Martine Chauvet, 13 Claude-Eric Bulabois, 14 Pascale Cony-Makhoul, 15 Selim Corm, 16 Jean-Yves Cahn, 1,14 and Pascal Mossuz …”

Myeloproliferative neoplasms in children

“…Similar to other MPNs, ET shows increased in vitro sensitivity to cytokines with increased colony formation (i.e., EEC growth), megakaryocytic progenitor sensitivity to TPO and endogenous megakaryocyte colony formation[20, 49]. …”