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Gossypol induces apoptosis in multiple myeloma cells by inhibition of interleukin-6 signaling and Bcl-2/Mcl-1 pathway.

Sadahira K, Sagawa M, Nakazato T, Uchida H, Ikeda Y, Okamoto S, Nakajima H, Kizaki M - Int. J. Oncol. (2014)

Bottom Line: In addition, Mcl-1 was downregulated by Gos.Moreover, JAK2 inhibition mimicked the effect of Gos in OPM2 cells including Bcl-2 dephosphorylation and Mcl-1 downregulation.These results demonstrated that Gos induces apoptosis in MM cells not only through displacing BH3-only proteins from Bcl-2, but also through inhibiting IL-6 signaling, which leads to Bcl-2 dephosphorylation and Mcl-1 downregulation.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology, Department of Internal Medicine, Keio University School of Medicine, Tokyo 160-8582, Japan.

ABSTRACT
Multiple myeloma (MM) is a clonal plasma cell disorder affecting the immune system with various systemic symptoms. MM remains incurable even with high dose chemotherapy using conventional drugs, thus necessitating development of novel therapeutic strategies. Gossypol (Gos) is a natural polyphenolic compound extracted from cotton plants, and has been shown to possess anti-neoplastic activity against various tumors. Recent studies have shown that Gos is an inhibitor for Bcl-2 or Bcl-XL acting as BH3 mimetics that interfere interaction between pro-apoptotic BH3-only proteins and Bcl-2/Bcl-XL. Since most of the patients with MM overexpress Bcl-2 protein, we considered Gos might be a promising therapeutic agent for MM. We herein show that Gos efficiently induced apoptosis and inhibited proliferation of the OPM2 MM cell line, in a dose- and time-dependent manner. Gos induced activation of caspase-3 and cytochrome c release from mitochondria, showing mitochondrial dysfunction pathway is operational during apoptosis. Further investigation revealed that phosphorylation of Bcl-2 at serine-70 was attenuated by Gos treatment, while protein levels were not affected. In addition, Mcl-1 was downregulated by Gos. Interestingly, phosphorylation of JAK2, STAT3, ERK1/2 and p38MAPK was inhibited by Gos-treatment, indicating that Gos globally suppressed interleukin-6 (IL-6) signals. Moreover, JAK2 inhibition mimicked the effect of Gos in OPM2 cells including Bcl-2 dephosphorylation and Mcl-1 downregulation. These results demonstrated that Gos induces apoptosis in MM cells not only through displacing BH3-only proteins from Bcl-2, but also through inhibiting IL-6 signaling, which leads to Bcl-2 dephosphorylation and Mcl-1 downregulation.

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Cell cycle analyses of OPM2 cells treated with Gossypol. (A) Cell cycle analysis of OPM2 cells with Gos. Cell were cultured with 5 μM of Gos for 0–24 h and then stained with PI as described in Materials and methods. DNA content was analyzed by flow cytometry. Representative figures from three independent experiments are shown. (B) Expression of cell cycle related proteins in OPM2 cells treated with 5 μM of Gos for the indicated times. β-actin was used as a loading control.
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f2-ijo-45-06-2278: Cell cycle analyses of OPM2 cells treated with Gossypol. (A) Cell cycle analysis of OPM2 cells with Gos. Cell were cultured with 5 μM of Gos for 0–24 h and then stained with PI as described in Materials and methods. DNA content was analyzed by flow cytometry. Representative figures from three independent experiments are shown. (B) Expression of cell cycle related proteins in OPM2 cells treated with 5 μM of Gos for the indicated times. β-actin was used as a loading control.

Mentions: Next we examined the effect of Gos on the cell cycle status of OPM2 cells. Cells were incubated with Gos and the cell cycle status was examined by flow cytometry at various time-points (Fig. 2A). The results demonstrated that Gos induced depletion of cells in S/G2/M phase and dramatic increase of cells in sub-G1 after 24 h of treatment. These results suggest that Gos induced cell cycle arrest at G1 followed by apoptosis in OPM2 cells.


Gossypol induces apoptosis in multiple myeloma cells by inhibition of interleukin-6 signaling and Bcl-2/Mcl-1 pathway.

Sadahira K, Sagawa M, Nakazato T, Uchida H, Ikeda Y, Okamoto S, Nakajima H, Kizaki M - Int. J. Oncol. (2014)

Cell cycle analyses of OPM2 cells treated with Gossypol. (A) Cell cycle analysis of OPM2 cells with Gos. Cell were cultured with 5 μM of Gos for 0–24 h and then stained with PI as described in Materials and methods. DNA content was analyzed by flow cytometry. Representative figures from three independent experiments are shown. (B) Expression of cell cycle related proteins in OPM2 cells treated with 5 μM of Gos for the indicated times. β-actin was used as a loading control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215583&req=5

f2-ijo-45-06-2278: Cell cycle analyses of OPM2 cells treated with Gossypol. (A) Cell cycle analysis of OPM2 cells with Gos. Cell were cultured with 5 μM of Gos for 0–24 h and then stained with PI as described in Materials and methods. DNA content was analyzed by flow cytometry. Representative figures from three independent experiments are shown. (B) Expression of cell cycle related proteins in OPM2 cells treated with 5 μM of Gos for the indicated times. β-actin was used as a loading control.
Mentions: Next we examined the effect of Gos on the cell cycle status of OPM2 cells. Cells were incubated with Gos and the cell cycle status was examined by flow cytometry at various time-points (Fig. 2A). The results demonstrated that Gos induced depletion of cells in S/G2/M phase and dramatic increase of cells in sub-G1 after 24 h of treatment. These results suggest that Gos induced cell cycle arrest at G1 followed by apoptosis in OPM2 cells.

Bottom Line: In addition, Mcl-1 was downregulated by Gos.Moreover, JAK2 inhibition mimicked the effect of Gos in OPM2 cells including Bcl-2 dephosphorylation and Mcl-1 downregulation.These results demonstrated that Gos induces apoptosis in MM cells not only through displacing BH3-only proteins from Bcl-2, but also through inhibiting IL-6 signaling, which leads to Bcl-2 dephosphorylation and Mcl-1 downregulation.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology, Department of Internal Medicine, Keio University School of Medicine, Tokyo 160-8582, Japan.

ABSTRACT
Multiple myeloma (MM) is a clonal plasma cell disorder affecting the immune system with various systemic symptoms. MM remains incurable even with high dose chemotherapy using conventional drugs, thus necessitating development of novel therapeutic strategies. Gossypol (Gos) is a natural polyphenolic compound extracted from cotton plants, and has been shown to possess anti-neoplastic activity against various tumors. Recent studies have shown that Gos is an inhibitor for Bcl-2 or Bcl-XL acting as BH3 mimetics that interfere interaction between pro-apoptotic BH3-only proteins and Bcl-2/Bcl-XL. Since most of the patients with MM overexpress Bcl-2 protein, we considered Gos might be a promising therapeutic agent for MM. We herein show that Gos efficiently induced apoptosis and inhibited proliferation of the OPM2 MM cell line, in a dose- and time-dependent manner. Gos induced activation of caspase-3 and cytochrome c release from mitochondria, showing mitochondrial dysfunction pathway is operational during apoptosis. Further investigation revealed that phosphorylation of Bcl-2 at serine-70 was attenuated by Gos treatment, while protein levels were not affected. In addition, Mcl-1 was downregulated by Gos. Interestingly, phosphorylation of JAK2, STAT3, ERK1/2 and p38MAPK was inhibited by Gos-treatment, indicating that Gos globally suppressed interleukin-6 (IL-6) signals. Moreover, JAK2 inhibition mimicked the effect of Gos in OPM2 cells including Bcl-2 dephosphorylation and Mcl-1 downregulation. These results demonstrated that Gos induces apoptosis in MM cells not only through displacing BH3-only proteins from Bcl-2, but also through inhibiting IL-6 signaling, which leads to Bcl-2 dephosphorylation and Mcl-1 downregulation.

Show MeSH
Related in: MedlinePlus