Interleukin-4 downregulates nuclear factor-erythroid 2 (NF-E2) expression in primary megakaryocytes and in megakaryoblastic cell lines

The trascriptional factor nuclear factor-erythroid 2 (NF-E2) is one of the few transcription factors known to be functionally linked to the megakaryoc ytic lineage, where it regulates terminal megakaryocyte maturation and plat elet formation, However, the regulation of NF-E2 expression in megakaryocyt ic cells has not been extensively evaluated, In particular, no data have be en reported on the effect of negative regulators of megakaryocytopoiesis on NF-E2 expression. This study investigated the in vitro effects of two nega tive regulators of megakaryocytopoiesis, such as interleukin-4 (IL-4) and t ransforming growth factor-beta1 (TGF-beta1) on the expression of NF-E2 tran scription factor in megakaryoblastic cell lines (Hel and MK1) and in normal CD34-derived megakaryocytic cells. For this purpose, we used quantitative real-time reverse transcription-polymerase chain reaction CRT-PCR) to detec t mRNA NF-E2 isoforms (a and f) and flow-cytometry analysis to evaluate NF- E2 protein expression. Our results demonstrated that TGF-beta1 did not inhi bit NF-E2 mRNA and protein expression of either maturating or fully mature normal megakaryocytic cells as well as that of the two cell lines. By contr ast, IL-4 downmodulates the expression of NF-E2, transcription factor at bo th mRNA and protein levels in normal maturating megakaryocytic cells and in the megakaryoblastic cell lines. NF-E2 expression of normal mature megakar yocytes was not affected by IL-4, Thus, the results of the present investig ation demonstrate that NF-E2 transcription factor is involved not only in t erminal megakaryocyte maturation but also in the negative regulation of the early phase of megakaryocyte development.