Korean J Hematol.  2005 Jun;40(2):93-100. 10.5045/kjh.2005.40.2.93.

Arsenic Trioxide Induces Erythroid Differentiation and Apoptosis of K562 Human Leukemia Cells through the Down-Regulation of Bcl-2

• You YK
• Cheong HJ
• Won JH
• Kim SJ
• Bae SB
• Kim CK
• Lee NS
• Lee KT
• Park SK
• Hong DS
• Park HS

• KMID: 2252343
• DOI: http://doi.org/10.5045/kjh.2005.40.2.93

Abstract

BACKGROUND
Arsenic trioxide (As2O3) has been identified as an effective drug for the treatment of acute promyelocytic leukemia (APL). However, the role of As2O3 during the erythroid differentiation of human leukemic cells remains unknown. In this study, we investigated the in vitro effects of As2O3 on the erythroid differentiation of the K562 cell line and also on the expression and regulation of the apoptotic modulators of this process.
METHODS
The K562 cells were cultured in the presence of 0.1, 0.5 and 1.0micrometer As2O3, or they were cultured in the presence of 1.0 and 10micrometer all trans retinoic acid (ATRA). The expression of glycophorin A before and after treatment with As2O3 or with ATRA in the K562 cells was assessed by flow cytometry and western blotting. The expressions of Bcl-2 and caspase-3 were determined by western blotting.
RESULTS
The viability of the K562 cells was not decreased after treating with 0.1 and 0.5micrometer of As2O3, but the viability was significantly reduced at a dose of 1.0micrometer Caspase 3 activation was not observed at 0.1 and 0.5micrometer of As2O3 until 12 days, but Caspase 3 was activated by 1.0micrometer of As2O3 from day 3. The expression of glycophorin A was increased in dose dependent manner by As2O3 treatment, but this was not changed in the ATRA treated K562 cells. The expression of Bcl-2 was increased by 0.1 and 0.5micrometer of As2O3, but it was abruptly reduced by 1.0micrometer of As2O3.
CONCLUSION
These results suggest that As2O3 induces the erythroid differentiation of K562 cells and that 1.0micrometer of As2O3 induces apoptosis through the down-regulation of Bcl-2.

Keyword

Arsenic trioxide; Differentiation; Apoptosis; ATRA; K562

MeSH Terms

Apoptosis*
Arsenic*
Blotting, Western
Caspase 3
Cell Line
Down-Regulation*
Flow Cytometry
Glycophorin
Humans*
K562 Cells
Leukemia*
Leukemia, Promyelocytic, Acute
Tretinoin
Arsenic
Caspase 3
Glycophorin
Tretinoin

Figure

• 

  Fig. 1. Viability of K562 cells after treatment with As2O 3 or ATRA. (A) The percentage of viable K562 cells after treatment with 1μM of As2O 3 began to decrease significantly 9 days after treatment. (B) The viabilities of K562 cells were 97%, 91%, and 74.5% by day 12 of treatment with 0.1μM, 0.5μM, and 1μM of As2O 3, respectively (∗P＜0.05, †P＜ 0.05). The viability of K562 cells was not changed by adding 1μM and 10μM of ATRA until day 12.

• 

  Fig. 2. C aspase 3 activation in K562 cells after treatment with As2O 3. Activation of cleaved caspase-3 (17kDa) was observed in K562 cells treated with 1μM of As2O3. Caspase-3 was not activated in K562 cells treated with a low dose As2O 3 (0.1μM and 0.5μM).

• 

  Fig. 3. Expression of glycophorin A in K562 cells after treatment with As2O 3 or ATRA by using the flow cytometry and western blot analysis. (A) Flow cytometric analysis showed that the intensity of glycophorin A expression was increased in a dose dependent fashion in K562 cells after 3 days incubation with As2O 3 but was unchanged in K562 cells incubated with ATRA. (B) Western blot analysis showed that treatment with As2O 3 induced upregulation of glycophorin A in a dose dependent fashion, whereas glycophorin A remained unaltered when treated with ATRA.

• 

  Fig. 4. Expression of Bcl-2 in K562 cells after treatment with As2O 3 or ATRA. The expression of Bcl-2 was increased in a time and dose dependent fashion in K562 cells treated with 0.1μM or 0.5μM of As2O 3 but abruptly decreased in K562 cells from 9 days after incubation with 1.0μM of As2O 3. Bcl-2 expression was not changed in K562 cells incubated with ATRA.

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