All-trans-retinoic acid effects the growth, differentiation and apoptosis of normal human myeloid progenitors derived from purified CD34+ bone marrow cells

Abstract

We have previously shown that all-trans retinoic acid (ATRA) increases the number of CFU-GM colonies grown from unseparated human bone marrow cells with crude sources of colony stimulating factors. In this study, we further characterized the effect of ATRA on the growth of CFU-GM stimulated by individual cytokines from multiple samples of CD34 ؉ enriched or purified human bone marrow cells. The number of IL-3-or GM-CSF-induced CFU-GM with 3 ؋ 10 ؊7 M ATRA was 3.25 ± 1.13, and 2.17 ؎ 0.8-fold greater respectively, compared to controls without ATRA, while G-CSF had no effect and the ratio of colony-induced with or without ATRA was 1.06 ؎ 0.17 (P ‫؍‬ 0.00012). No colonies grew with ATRA ؉ IL-6 or ATRA without a cytokine. Maximum enhancing effect on IL-3-induced CFU-GM occurred when ATRA was added on day 2, gradually diminished when delaying ATRA, and in cultures of day 9 or older adding ATRA had no effect. In 14 days liquid cultures of purified CD34 ؉ cells with IL-3, ATRA increased the number of myeloid differentiated cells to 91-95%, compared to 37-70% with IL-3 alone. In addition, the number of apoptotic cells using the annexin V method increased after 14 days from 5.1% with IL-3 to 17.1% with IL-3 ؉ ATRA and by the TUNEL in situ method from 10-26% to 60-95%, respectively. This study demonstrates that ATRA consistently enhances the growth of myeloid progenitors from CD34 ؉ cells. This effect is dependent on the stimulating cytokine, suggesting the myeloid cells responding to ATRA are the less mature CFU-GMs that are targets of IL-3 and GM-CSF and not the G-CSF-responding mature progenitors. The growth stimulation by ATRA and IL-3 is also associated with granulocyte differentiation and increased apoptosis. These studies further suggest a potential role of pharmacological doses of ATRA on the development of normal human hematopoietic cells. Leukemia (2000) 14, 874-881.

The role of retinoids and retinoic acid receptors in normal hematopoiesis

“…14,31,32 Since G-CSF-dependent colonies likely arise from more mature progenitors than either IL-3-or GM-CSF-dependent colonies, this suggests that ATRA may regulate the proliferation/differentiation of cells at a relatively early stage of myeloid maturation. 33 …”

The cytokines IL-3 and GM-CSF regulate the transcriptional activity of retinoic acid receptors in different in vitro models of myeloid differentiation

“…Knockout mice deficient in both RAR␣ and RAR␥ display an in vitro block to granulocyte differentiation, 3 and RA stimulates the growth and differentiation of granulocyte progenitors in cytokine-stimulated cultures of purified CD34 ϩ cells. 4 Importantly, the 15;17 chromosome translocation in human acute promyelocytic leukemia (APL) generates the dominant negative PML-RAR␣ fusion protein that inhibits the function of normal RARs, [5][6][7] resulting in the block to terminal granulocytic differentiation that characterizes this leukemia.…”

Signaling revisited in acute promyelocytic leukemia

“…RARs are retinoic acid (RA)-dependent transcription factors 41 involved in the control of hematopoietic cells differentiation. 42,43 Interestingly, PML, PLZF and STAT5b are involved in the control of transduction of the cell mitogenic and survival signals. [44][45][46][47] In more than 95% of APL, the specific translocation t (15;17) 48 creates a PML-RAR␣ and reciprocal fusion proteins.…”