Developmental expression of plasminogen activator inhibitor-1 associated with thrombopoietin-dependent megakaryocytic differentiation

Plasminogen activator inhibitor-1 (PAI-1) is present in the platelet a-gran ule and is released on activation. However, there is some debate as to whet her the megakaryocyte and platelet synthesize PAI-1, take it up from plasma , or both. We examined the expression of PAI-1 in differentiating megakaryo cytic progenitor cells (UT-7) and in CD34(+)/CD41(-) cells from cord blood. UT-7 cells differentiated with thrombopoietin (TPO) resembled megakaryocyt es (UT-7/TPO) with respect to morphology, ploidy, and the expression of gly coprotein IIb-IIIa. PAI-1 messenger RNA (mRNA) expression was upregulated a nd PAI-1 protein synthesized in the UT-7/TPO cells accumulated in the cytop lasm without being released spontaneously. In contrast, erythropoietin (EPO )-stimulated UT-7 cells (UT-7/EPO) did not express PAI-1 mRNA after stimula tion with TPO because they do not have endogenous c-Mpl. After cotransfecti on with human wild-type c-mpl, the cells (UT-7/EPO-MPL) responded to phorbo l 12-myristate 13-acetate (PMA), tumor necrosis factor-alpha (TNF-alpha), a nd interleukin-1 beta (IL-1 beta) with enhanced PAI-1 mRNA expression withi n 24 to 48 hours. However, induction of PAI-1 mRNA in UT-7/EPO-MPL cells by TPO required at least 14-days stimulation. UT-7/EPO cells expressing c-Mpl changed their morphology and the other characteristics similar to the UT-7 /TPO cells, TPO also differentiated human cord blood CD34(+)/CD41(-) cells to CD34(-)/CD41(+) cells, generated morphologically mature megakaryocytes, and induced the expression of PAI-1 mRNA, These results suggest that both P AI-1 mRNA and de novo PAI-1 protein synthesis is induced after differentiat ion of immature progenitor cells into megakaryocytes by TPO, (C) 1999 by Th e American Society of Hematology.