Amiloride suppresses erythropoietin-induced proliferation and MAP kinase, but potentiates differentiation of J2E cells.
The J2E erythroid cell line proliferates and differentiates in response to erythropoietin (epo). Here we demonstrate that the diuretic amiloride can suppress normal and hormone-induced cell division in a dose-dependent manner. In the presence of amiloride, cell numbers did not increase, [3H]thymidine incorporation decreased, and fewer cells were observed in the S, G2, and M phases of the cell cycle. In addition, the levels of proliferating cell nuclear antigen, a subunit of DNA polymerase delta, fell. In marked contrast, epo-initiated differentiation was potentiated when J2E cells were cultured with the drug: the number of benzidine-positive cells increased, hemoglobin content per cell rose, and more morphologically mature cells were produced. Immunoblotting with anti-phosphotyrosine antibodies revealed that amiloride reduced the number of phosphorylated proteins in epo-stimulated cells. Moreover, the protein content of p42 and p44 MAP kinases was noticeably downregulated in amiloride-treated cultures. These data indicate that amiloride may interfere with epo-induced signaling cascades within J2E cells which result in restricted cell division and promotion of maturation.
Callus
B
B
Department of Biochemistry, University of Western Australia, Nedlands.
Tilbrook
P A
PA
Busfield
S J
SJ
Cull
V S
VS
Bittorf
T
T
Klinken
S P
SP
D000584:Amiloride / Q000494:pharmacology*; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects*; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004912:Erythrocytes; D004921:Erythropoietin / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D019833:Genistein; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007529:Isoflavones / Q000494:pharmacology; D007700:Kinetics; D007930:Leucine / Q000378:metabolism; D010750:Phosphoproteins / Q000032:analysis / Q000378:metabolism; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011994:Recombinant Proteins / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D014443:Tyrosine / Q000031:analogs & derivatives / Q000032:analysis
D006454:Hemoglobins; D007529:Isoflavones; D010750:Phosphoproteins; D011994:Recombinant Proteins; D004921:Erythropoietin; D019000:Phosphotyrosine; D014443:Tyrosine; D000584:Amiloride; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D007930:Leucine; D013936:Thymidine