Overexpression of nuclear distribution protein (hNUDC) causes pro-apoptosis and differentiation in Dami megakaryocytes.

OBJECTIVES: Overexpression of hNUDC, a member of the nuclear distribution protein family, reduces cell population growth in prostate cancer cell lines, concurrent with induced morphological change and enhanced polyploidization. These phenomena are also closely associated with terminal phases of megakaryocyte maturation.
MATERIALS AND METHODS: In Dami cells, MTT and trypan blue assays were used to investigate cell viability and proliferation effects of hNUDC, and flow cytometry was used to analyse cell cycle and DNA content. Real-time RT-PCR was employed to detect mRNA expression. Activations of caspase-3, ERK, Akt and Stat-5 were determined by immunoblotting. May-Grünwald-Giemsa staining was performed to reveal cell morphology. RESULTS AND
CONCLUSION: Functional studies using adenovirus-mediated hNUDC overexpression led to inhibition of megakaryocyte proliferation via cell cycle arrest in G2/M transition phase. This process could have been be mediated by upregulation of p21 and downregulation of its downstream targets, including cyclin B1, cyclin B2 and c-myc. Enhanced apoptosis in turn ensued, characterized by increased caspase-3 activation, upregulation of pro-apoptotic Bax and downregulation of anti-apoptotic Bcl-2. Furthermore, hNUDC overexpression elevated the level of megakaryocyte maturation, associated with increased polyploidy, cell morphological changes and increased expression of cell surface differentiation markers, including CD10, CD44, CD41 and CD61. Our results further suggest that the ERK signalling pathway was involved in hNUDC overexpression-induced apoptosis. Taken together, this study provides experimental evidence for overexpression of hNUDC in Dami cells and suggests that activation of apoptotic machinery may be involved in megakaryocytic differentiation.