Blood, Vol. 91 No. 2 (January 15), 1998:
pp. 724-725
CORRESPONDENCE
A Retinoic Acid Receptor-
(RAR) Selective Agonist
Modulates Procoagulant Activity of Acute Promyelocytic Cells and
Induces Their Differentiation Into Neutrophils
 |
LETTER |
To the Editor:
We recently found that a subtype of retinoic acid receptors (RARs),
RAR
, mediates upregulation of TM gene expression in
human promyelocytic leukemia cells (NB4), acute monocytic leukemia
cells, and human umbilical vein endothelial cells and that the
cooperation of RAR and RAR
exerts downregulation of TF gene
expression in NB4 cells.1 We introduce here new data to
support our previous findings using RAR, RAR, and RAR
selective agonists, Ro40-6055, Ro48-2249, and Ro44-4753, respectively.
These synthetic retinoids were kindly provided by F. Hoffman-La Roche,
Ltd (Basel, Switzerland). Our experiments also showed that the
incubation of NB4 cells with the RAR selective agonist, Ro40-6055,
induced cell differentiation into neutrophils and apoptosis. These
effects were not exerted by Ro48-2249 and Ro44-4753.
When NB4 cells were incubated with 0.01 µmol/L of Ro40-6055,
Ro48-2249, or Ro44-4753 for 24 hours, Ro40-6055 caused induction of TM
antigen and reduction of TF antigen (Fig 1A and
B), which was similar to the results
induced by a synthetic retinoid Am80, which is an agonist to RAR and
RAR.1 The TM upregulation by the incubation with
Ro40-6055 was more effective than that by the stimulation of the
combination with Ro40-6055 and Ro48-2249 (Fig 1A). Ro40-6055 and
Ro48-2249 showed downregulation of TF in NB4 cells, and the TF
downregulation by the combined incubation of Ro40-6055 and Ro48-2249
was more significant than that by each retinoid (Fig 1B).
Interestingly, the incubation of NB4 cells with Ro44-4753 induced
upregulation of TF antigen level, although its mechanism is not yet
known.
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| Fig 1.
Effects of RAR, RAR, and RAR selective retinoids
on the expression of the toal TM an TF antigens in NB4 cells. NB4 cells were incubated with 0.01 µmol/L of Ro40-6055 (lane 2), Ro48-2249 (lane 3), Ro44-4753 (lane 4), or a combination of 0.005 µmol/L each
of Ro40-6055 and Ro48-2249 (lane 5) for 24 hours. Total TM (A) and TF
(B) antigen levels in cell lysates were measured by enzyme-linked
immunosorbent assay, as previously reported.1 Results are
shown as the means ± SD of three different experiments.
|
|
NB4 cells were further incubated with 0.01 µmol/L of Ro40-6055 for 7 days and were evaluated for changes in the morphology and surface
markers of the leukemic cells. NB4 cells with frequent mitotic figures
and morphologic features characteristic of acute promyelocytic leukemia
cells were shown to have significant morphologic changes, such as
condensed chromatin, smaller nuclei, decreased nuclei/cytoplasm ratio,
and appearance of neutrophilic granules in the cytoplasm when treated
with Ro40-6055. Differentiation of NB4 cells was also assessed by flow
cytometric analysis for several membrane-bound differentiation markers.
Expression of the cell surface antigens CD11b (a marker of -integrin
subunit expressed by both granulocytes and monocytes), CD11c
(granulocytic lineage), and CD14 (late monocytic cell) is gradually
detected during the normal development of mature monocytes from
hematopoietic stem cells.2 The expression levels of CD11b
and CD11c antigens (19.1% and 43.2%, respectively) were significantly
increased after incubation with 0.01 µmol/L of Ro40-6055 to 91.6%
and 90.8%, respectively. CD14 expression level was only slightly
increased from 6.5% to 8.4%. DNA fragmentation assay of NB4 cells
showed the occurrence of apoptosis by Ro40-6055 stimulation (0.01 µmol/L for 7 days).
These results support the idea that RAR mediates TM gene expression
and that RAR and also RAR are relevant to the retinoid-induced TF
downregulation. A selective RAR agonist, Ro40-6055, is a potent inducer of differentiation and apoptosis with anticoagulant effect on
acute promyelocytic leukemia cells.
Misako Shibakura
Takatoshi Koyama
Mai Ohsawa
Ryuichi Kamiyama
Shinsaku Hirosawa
The School of Allied Health Sciences
First Department
of Internal Medicine
Tokyo Medical and Dental University
Tokyo,
Japan
| |
REFERENCES |
1.
Shibakura M,
Koyama T,
Saito T,
Miyasaka N,
Kamiyama R,
Hirosawa S:
Anticoagulant effects of synthetic retinoids mediated via different receptors on human leukemia and umbilical vein endothelial cells.
Blood
90:1545,
1997[Abstract/Free Full Text]
2.
Elstner E,
Linker-Israeli M,
Le J,
Umiel T,
Michl P,
Said JW,
Binderup L,
Reed JC,
Koeffler HP:
Synergistic decrease of clonal proliferation, induction of differentiation, and apoptosis of acute promyelocytic leukemia cells after combined treatment with novel 20-epi vitamin D3 analogs and 9-cis retinoic acid.
J Clin Invest
99:349,
1997[Medline]
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