Recombinant human interleukin-3 and granulocyte-macrophage colony-stimulating factor show common biological effects and binding characteristics on human monocytes.
Two human hemopoietic growth factors involved in monocytopoiesis, interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) were studied for their ability to stimulate blood monocytes and to bind to the monocyte membrane. Both cytokines maintained monocyte/macrophage numbers during long-term culture and increased cell size as compared with controls. Effects on cell numbers were present at low cytokine concentrations (6 to 20 pmol/L), whereas enhanced 3H-thymidine incorporation was observed only at higher concentrations (greater than or equal to 60 pmol/L). Autoradiographic studies showed only 1% to 3% of stimulated monocytes with nuclear grains. These results suggest that the primary mechanism for IL-3 and GM-CSF-induced maintenance of monocyte/macrophage numbers in humans is through an effect on cell survival. Surface receptors for both IL-3 and GM-CSF were studied by using 125I-labeled recombinant human (rh) cytokines and performing Scatchard analyses. Both cytokines showed curvilinear Scatchard plots, and computer analyses favored a two-site binding model. High-affinity binding data for 125I rhIL-3 (Kd 7.7 to 38.2 pmol/L; receptor number/cell 95 to 580) and for 125I rhGM-CSF (Kd 4.7 to 38.9 pmol/L; receptor number/cell 8 to 67) show similar binding affinities for the two cytokines but a lower receptor number/cell for 125I rhGM-CSF. Low-affinity binding characteristics for 125I rhIL-3 (Kd 513 to 939 pmol/L; receptor number/cell 179 to 5,274) and for 125I rhGM-CSF (Kd 576 to 1,120 pmol/L; receptor number/cell 130 to 657) show a similar pattern for the two cytokines. Specificity of 125I rhIL-3 and 125I rhGM-CSF binding to monocytes was established by the ability of the homologous cytokine to inhibit binding and the inability of a range of other cytokines to compete at 100-fold excess molar concentration. It is important, however, that binding of 125I rhIL-3 was partially inhibited by rhGM-CSF and that rhIL-3 partially inhibited binding of 125I rhGM-CSF to the monocyte membrane under conditions shown to prevent receptor internalization. The degree of inhibition varied between 25% and 80% in different experiments, and quantitative inhibition experiments showed that 1,000-fold excess concentrations of competitor failed to inhibit binding of the heterologous ligand completely. These results demonstrate that human IL-3 and GM-CSF have similar effects on growth and survival of human monocytes in vitro and suggest that these and other common biological effects may be mediated either through a common receptor or through distinct receptors associated on the monocyte membrane.
Elliott
M J
MJ
Division of Human Immunology, Royal Adelaide Hospital, South Australia.
Vadas
M A
MA
Eglinton
J M
JM
Park
L S
LS
To
L B
LB
Cleland
L G
LG
Clark
S C
SC
Lopez
A F
AF
D001667:Binding, Competitive; D002455:Cell Division / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D003115:Colony-Stimulating Factors / Q000378:metabolism / Q000494:pharmacology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D006133:Growth Substances / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D007377:Interleukin-3 / Q000378:metabolism / Q000494:pharmacology*; D007700:Kinetics; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism; D011994:Recombinant Proteins
D003115:Colony-Stimulating Factors; D006133:Growth Substances; D007377:Interleukin-3; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor