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Interleukin-4 Downregulates Nuclear Factor-Erythroid 2 (NF-E2) Expression in Primary Megakaryocytes and in Megakaryoblastic Cell Lines

Istituto di Ematologia e Oncologia Medica "L. e A. Seràgnoli;" University of Bologna-Italy

Key Words. NF-E2 • IL-4 • TGF-ß1 • Megakaryocytes • Real-time RT-PCR

Lucia Catani, Ph.D., Istituto di Ematologia e Oncologia Medica, "L. e A. Seràgnoli," Ospedale S. Orsola, Via Massarenti 9-40138, Bologna, Italy. Telephone: 051-636-4038; Fax: 051-636-4037; e-mail: lcatani{at}med.unibo.it

The trascriptional factor nuclear factor-erythroid 2 (NF-E2) is one of the few transcription factors known to be functionally linked to the megakaryocytic lineage, where it regulates terminal megakaryocyte maturation and platelet formation. However, the regulation of NF-E2 expression in megakaryocytic cells has not been extensively evaluated. In particular, no data have been reported on the effect of negative regulators of megakaryocytopoiesis on NF-E2 expression. This study investigated the in vitro effects of two negative regulators of megakaryocytopoiesis, such as interleukin-4 (IL-4) and transforming growth factor-ß1 (TGF-ß1) on the expression of NF-E2 transcription 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 (RT-PCR) to detect mRNA NF-E2 isoforms (a and f) and flow-cytometry analysis to evaluate NF-E2 protein expression. Our results demonstrated that TGF-ß1 did not inhibit 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 contrast, IL-4 downmodulates the expression of NF-E2 transcription factor at both mRNA and protein levels in normal maturating megakaryocytic cells and in the megakaryoblastic cell lines. NF-E2 expression of normal mature megakaryocytes was not affected by IL-4. Thus, the results of the present investigation demonstrate that NF-E2 transcription factor is involved not only in terminal megakaryocyte maturation but also in the negative regulation of the early phase of megakaryocyte development.