title: Abolished adherence alters signaling pathways in phorbol ester-induced human U937 cells.
abstract: Phorbol ester (TPA) treatment of human U937 myeloid leukemia cells is associated with increasing adherence and monocyte-like maturation whereby the role of β2 integrin-mediated attachment for subsequent growth properties and the differentiation program remains unclear. Here, stably-transfected U937 cells with a pMTH1 vector containing the β2 integrin gene of CD11b in antisense orientation (asCD11b-U937) demonstrated a significantly reduced proliferative capacity in contrast to control vector transfectants (pMTH1-U937) or wild-type U937 cells. Phorbol ester exposure induced adherence and growth arrest in more than 90% of pMTH1-U937 and wild-type U937 cells after 72 h. In contrast, TPA-treated asCD11b-U937 failed to attach and the proliferation continued in more than 30% of the cells. Moreover, increased apoptosis appeared in asCD11b-U937 after TPA induction in contrast to pMTH1-U937 cells. In addition, non-specific inhibition of adherence on an agarose surface demonstrated internucleosomal DNA fragmentation in both, pMTH1-U937 and asCD11b-U937 after TPA treatment indicating a functional relationship between abolished adherence, regulation of proliferation and induction of apoptosis. Western blot analysis revealed differences in the expression levels and altered phosphorylation patterns of Pyk-2, pp60src and p42/p44 MAP kinases between pMTH1-U937 and asCD11b-U937 following TPA exposure which was also substantiated by Pyk-2 immunoprecipitation. These findings suggested that induced adherence predominantly mediated by a functional CD11b/CD18 integrin in U937 cells is involved in the activation of downstream signaling kinases and contributes to cell cycle regulation and apoptosis during monocytic maturation.
authors.affiliation: Biochemistry and Tumor Biology Lab, Gynecology Research Unit, Dept, of Gynecology and Obstetrics, Medical University, Hannover, Germany. hass.ralf@mh-hannover.de.
authors.forename: Anna
authors.initials: A
authors.lastname: Otte
authors.forename: Katharina
authors.initials: K
authors.lastname: Mandel
authors.forename: Gesche
authors.initials: G
authors.lastname: Reinstrom
authors.forename: Ralf
authors.initials: R
authors.lastname: Hass