Mechanisms mediating the inhibitory effect of all-trans retinoic acid on primitive hematopoietic stem cells in human long-term bone marrow culture.

All-trans retinoic acid (RA) has generally been found to stimulate late committed (colony-forming unit- granulocyte, macrophage [CFU-GM]) and inhibit early (CFU-Blast) normal human myeloid progenitor cells. The present study provides the first evidence that the pharmacological concentration of 1 microM RA, exerts an inhibitory effect on the proliferation of functional human primitive hemopoietic stem cells (cobblestone area-forming cell [CFAC]) in long-term bone marrow cultures. Treatment of four-week confluent bone marrow culture with 1 microM RA for five days significantly reduced week 4 CAFC from 88 +/- 10 in control cultures to only 52 +/- 12 per 10(5) cells, p < 0.01. Quantitative enzyme-linked immunosorbent assay measurement of interleukin 6 (IL-6) and IL-11 produced from the four-week bone marrow stroma culture revealed only a slight and moderate increase of IL-6 and IL-11 production after treatment with RA. On the other hand, treatment with RA profoundly increased the soluble receptor gp130 released from the four-week bone marrow stroma by 7.5-fold from only 145 +/- 2.1 pg per ml in control cultures to 1,069.9 +/- 3.8 pg per ml in RA-treated cultures. A similar marked increase in the soluble adhesion molecules ICAM-1, and to a lesser extent VCAM-1, released from the four-week bone marrow stroma was observed after RA treatment. IL-6 has been implicated in the inhibitory effect of RA in several human hemopoietic and nonhemopoietic cells. The common transducing signal chain gp130, for all receptors of the IL-6 cytokine family, is expressed in most primitive human hemopoietic CD34(+) cells and its signaling was shown to synergize with other hemopoietic cytokines to expand primitive human hemopoietic stem cells. Recently, soluble gp130 was shown to be a natural potent antagonist of the human IL-6 cytokine family by binding the ligand and thereby reducing its bioavailability. The profound and rapid 7.5-fold increase in the natural antagonist of human IL-6 cytokine family after RA treatment could abrogate the gp130 signaling required for proliferation and/or expansion of human primitive hemopoietic stem cells and lead to the observed inhibitory effect of RA on CAFC. Both adhesion molecules VCAM-1 and ICAM-1 mediate human hemopoietic stem cell adhesion to marrow stroma. The present significant increase in the soluble form of these adhesion molecules after RA treatment could exert a significant antagonist effect on their function and hence may impair CAFC adhesion to marrow stroma. In conclusion, the RA inhibitory effect on the proliferation of primitive human hemopoietic stem cells could be mediated through: A) an impaired hemopoietic stem cell adhesion due to the significant increase in soluble adhesion molecules released from the marrow stroma after RA treatment, and B) a significantly reduced gp130 signaling that is necessary for stem cell proliferation due to the natural antagonistic effect of the profoundly increased level of soluble gp130 released from the marrow stroma after treatment with RA.