UI  - 94129004
TI  - Retinoic acid downmodulates erythroid differentiation and GATA1 expression in purified adult-progenitor culture.
AB  - All-trans retinoic acid (RA) is an important morphogen in vertebrate development, a normal constituent in human adult blood and is also involved in the control of cell growth and differentiation in acute promyelocytic leukemia.
We have examined the effects of RA on normal hematopoiesis by using early hematopoietic progenitor cells (HPC) stringently purified from adult peripheral blood.
In clonogenetic fetal calf serum-supplemented (FCS+) or -nonsupplemented (FCS-) culture treated with saturating levels of interleukin-3 (IL-3) granulocyte- macrophage colony-stimulating factor (GM-CSF) and erythropoietin (Ep) (combined with c-kit ligand in FCS(-)-culture conditions), RA induces a dramatic dose-dependent shift from erythroid to granulomonocytic colony formation, the latter colonies being essentially represented by granulocytic clones.
This shift is apparently not caused by a recruitment phenomenon, because in FCS+ culture, the total number of colonies is not significantly modified by RA addition.
In FCS- liquid- suspension culture supplemented with saturating Ep level and low-dose IL-3/GM-CSF, adult HPC undergo unilineage erythropoietic differentiation: Here again, treatment with high-dose RA induces a shift from the erythroid to granulocytic differentiation pathway. Studies on RA time-response or pulse treatment in semisolid or liquid culture show that early RA addition is most effective, thus indicating that early but not late HPC are sensitive to its action.
We then analyzed the expression of the master GATA1 gene, which encodes a finger transcription factor required for normal erythroid development; addition of RA to HPC stimulated into unilineage erythropoietic differentiation in liquid culture caused a virtually complete inhibition of GATA1 mRNA induction.
These results indicate that RA directly inhibits the erythroid differentiation program at the level of early adult HPC, and may lead to a shift from the erythroid to granulocytic differentiation pathway.
This phenomenon is correlated with inhibition of GATA1 induction in the early stages of erythropoietic differentiation.

UI  - 94134077
TI  - Induction of phosphatidylinositol turnover and EGR-1 mRNA expression by crosslinking of surface IgM and IgD in the human B cell line B104.
AB  - We have previously shown that a human B lymphoma cell line, B104, expressed surface IgM (sIgM) and surface IgD (sIgD), and that crosslinking of sIgM and sIgD by anti-IgM antibody (Ab) and anti-IgD Ab, respectively, induced Ca2+ influx to almost the same degree, whereas only sIgM-crosslinking caused B104 cell death.
Here, we investigated the accumulation of cyclic AMP (cAMP), the hydrolysis of inositol phosphates, protein kinase C (PKC) activity and the induction of Egr-1 and c-fos mRNA expression by sIgM- and sIgD-crosslinking to examine differences in the signals mediated through sIgM and sIgD in B104 cells.
Both sIgM- and sIgD-crosslinking with antibodies induced elevation of cAMP levels, phosphatidylinositol turnover, PKC activation and expression of Egr-1 and c-fos mRNA, although sIgM-crosslinking was more effective than sIgD-crosslinking, presumably due to the higher expression of sIgM than of sIgD.
Egr-1 mRNA expression induced by sIgM- and sIgD-crosslinking was inhibited by H7, erbstatin and genistein, but not by HA1004.
Erbstatin and genistein inhibited the sIg-crosslinking- induced Egr-1 mRNA expression in a dose-dependent manner parallel to that observed in the inhibition of sIg-crosslinking-induced protein tyrosine phosphorylation.
Phorbol myristate acetate induced Egr-1 mRNA expression but forskolin and dibutyryl cyclic AMP did not.
These findings suggest that the Egr-1 mRNA activating signals through sIgM and sIgD are protein tyrosine kinase- and PKC-dependent, but protein kinase A-independent.
Cyclosporin A (CsA) and FK506 rescued B104 cells from death induced by anti-IgM Ab, but did not affect the expression of Egr-1 and c-fos mRNA, showing that CsA and FK506 affect signal transducers differently from or downstream to these molecules.
The difference in signals transduced through sIgM and sIgD in B104 cells is discussed.
