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Autocrine prostaglandin E₂ signaling promotes promonocytic leukemia cell survival via COX-2 expression and MAPK pathway.

Shehzad A, Lee J, Lee YS - BMB Rep (2015)

Bottom Line: Treatment with PGE₂ activated anti-apoptotic proteins such as Bcl-2 and Bcl-xL while reducing pro-apoptotic proteins, thereby enhancing cell survival.Silencing and inhibition of COX-2 with siRNA transfection or treatment with indomethacin led to a pronounced reduction of the extracellular levels of PGEv, and restored the menadione- induced cell death.These results demonstrate that PGE₂ signaling acts in an autocrine manner, and specific inhibition of PGE₂ will provide a novel approach for the treatment of leukemia.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences, BK21 Plus KNU Creative BioResearch Group, Kyungpook National University, Daegu 702-701, Korea; Department of Biomedical Engineering and Sciences, School of Mechanical & Manufacturing Engineering, National University of Sciences & Technology, Islamabad, Pakistan.

ABSTRACT
The COX-2/PGE₂ pathway has been implicated in the occurrence and progression of cancer. The underlying mechanisms facilitating the production of COX-2 and its mediator, PGE₂, in cancer survival remain unknown. Herein, we investigated PGE₂-induced COX-2 expression and signaling in HL-60 cells following menadione treatment. Treatment with PGE₂ activated anti-apoptotic proteins such as Bcl-2 and Bcl-xL while reducing pro-apoptotic proteins, thereby enhancing cell survival. PGE₂ not only induced COX-2 expression, but also prevented casapse-3, PARP, and lamin B cleavage. Silencing and inhibition of COX-2 with siRNA transfection or treatment with indomethacin led to a pronounced reduction of the extracellular levels of PGEv, and restored the menadione- induced cell death. In addition, pretreatment of cells with the MEK inhibitor PD98059 and the PKA inhibitor H89 abrogated the PGE₂-induced expression of COX-2, suggesting involvement of the MAPK and PKA pathways. These results demonstrate that PGE₂ signaling acts in an autocrine manner, and specific inhibition of PGE₂ will provide a novel approach for the treatment of leukemia.

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COX-2 siRNA demolishes PGE2- induced expression of COX-2. (A) Cells were exposed to different doses of PGE2 for 12 h and to 1 μM PGE2 for the indicated times. PGE2 induced COX-2 expression in a dose- and time- dependent manner. (B) PGE2 induced COX-2 expression via the MEK and PKA pathways. Cells were pretreated with 10 μM PD98059 or 1 μM H89 for 2 h, and then exposed to PGE2, which failed to induce COX-2 expression. (C) Cells were stably transfected with different concentrations of COX-2 siRNA and then exposed to PGE2. Transfection with COX-2 siRNA prevented PGE2-induced expression of COX-2 at a dose of 10 nM. Scrambled siRNA displayed similar effects as the control upon PGE2 treatment.
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Figure 003: COX-2 siRNA demolishes PGE2- induced expression of COX-2. (A) Cells were exposed to different doses of PGE2 for 12 h and to 1 μM PGE2 for the indicated times. PGE2 induced COX-2 expression in a dose- and time- dependent manner. (B) PGE2 induced COX-2 expression via the MEK and PKA pathways. Cells were pretreated with 10 μM PD98059 or 1 μM H89 for 2 h, and then exposed to PGE2, which failed to induce COX-2 expression. (C) Cells were stably transfected with different concentrations of COX-2 siRNA and then exposed to PGE2. Transfection with COX-2 siRNA prevented PGE2-induced expression of COX-2 at a dose of 10 nM. Scrambled siRNA displayed similar effects as the control upon PGE2 treatment.

Mentions: Several pharmacological and epidemiological studies have shown that induction of COX-2 correlates with tumorigenesis and poor prognosis (12). Therefore, the effect of PGE2 on COX-2 expression was next evaluated. As shown in Fig. 3A, time-dependent activation of COX-2 expression was observed, which suggests that PGE2 caused up-regulation of COX-2, at least in part, by autocrine signaling and provision of a positive feedback mechanism. As the MAPK pathway was shown to mediate the anti-apoptotic activity of PGE2, and we previously confirmed the involvement of PKA (7), inhibition of MEK and PKA was carried out with PD98059 and H89 inhibitors, respectively. Pretreatment of cells with PD98059 or H89 prevented the PGE2-induced expression of COX-2, suggesting roles of the MAPK and PKA pathways (Fig. 3B). Small interfering RNA (siRNA) was then utilized for functional gene silencing to study COX-2 genetic loss-of-function in HL-60 cells upon PGE2 treatment. Transfection of cells with 10 nM COX-2 siRNA diminished the PGE2-induced expression of COX-2 (Fig. 3C). These results suggest that PGE2 mediates COX-2 expression via the MAPK and PKA pathways, and may be processed in an autocrine manner.


Autocrine prostaglandin E₂ signaling promotes promonocytic leukemia cell survival via COX-2 expression and MAPK pathway.

Shehzad A, Lee J, Lee YS - BMB Rep (2015)

COX-2 siRNA demolishes PGE2- induced expression of COX-2. (A) Cells were exposed to different doses of PGE2 for 12 h and to 1 μM PGE2 for the indicated times. PGE2 induced COX-2 expression in a dose- and time- dependent manner. (B) PGE2 induced COX-2 expression via the MEK and PKA pathways. Cells were pretreated with 10 μM PD98059 or 1 μM H89 for 2 h, and then exposed to PGE2, which failed to induce COX-2 expression. (C) Cells were stably transfected with different concentrations of COX-2 siRNA and then exposed to PGE2. Transfection with COX-2 siRNA prevented PGE2-induced expression of COX-2 at a dose of 10 nM. Scrambled siRNA displayed similar effects as the control upon PGE2 treatment.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4352612&req=5

Figure 003: COX-2 siRNA demolishes PGE2- induced expression of COX-2. (A) Cells were exposed to different doses of PGE2 for 12 h and to 1 μM PGE2 for the indicated times. PGE2 induced COX-2 expression in a dose- and time- dependent manner. (B) PGE2 induced COX-2 expression via the MEK and PKA pathways. Cells were pretreated with 10 μM PD98059 or 1 μM H89 for 2 h, and then exposed to PGE2, which failed to induce COX-2 expression. (C) Cells were stably transfected with different concentrations of COX-2 siRNA and then exposed to PGE2. Transfection with COX-2 siRNA prevented PGE2-induced expression of COX-2 at a dose of 10 nM. Scrambled siRNA displayed similar effects as the control upon PGE2 treatment.
Mentions: Several pharmacological and epidemiological studies have shown that induction of COX-2 correlates with tumorigenesis and poor prognosis (12). Therefore, the effect of PGE2 on COX-2 expression was next evaluated. As shown in Fig. 3A, time-dependent activation of COX-2 expression was observed, which suggests that PGE2 caused up-regulation of COX-2, at least in part, by autocrine signaling and provision of a positive feedback mechanism. As the MAPK pathway was shown to mediate the anti-apoptotic activity of PGE2, and we previously confirmed the involvement of PKA (7), inhibition of MEK and PKA was carried out with PD98059 and H89 inhibitors, respectively. Pretreatment of cells with PD98059 or H89 prevented the PGE2-induced expression of COX-2, suggesting roles of the MAPK and PKA pathways (Fig. 3B). Small interfering RNA (siRNA) was then utilized for functional gene silencing to study COX-2 genetic loss-of-function in HL-60 cells upon PGE2 treatment. Transfection of cells with 10 nM COX-2 siRNA diminished the PGE2-induced expression of COX-2 (Fig. 3C). These results suggest that PGE2 mediates COX-2 expression via the MAPK and PKA pathways, and may be processed in an autocrine manner.

Bottom Line: Treatment with PGE₂ activated anti-apoptotic proteins such as Bcl-2 and Bcl-xL while reducing pro-apoptotic proteins, thereby enhancing cell survival.Silencing and inhibition of COX-2 with siRNA transfection or treatment with indomethacin led to a pronounced reduction of the extracellular levels of PGEv, and restored the menadione- induced cell death.These results demonstrate that PGE₂ signaling acts in an autocrine manner, and specific inhibition of PGE₂ will provide a novel approach for the treatment of leukemia.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences, BK21 Plus KNU Creative BioResearch Group, Kyungpook National University, Daegu 702-701, Korea; Department of Biomedical Engineering and Sciences, School of Mechanical & Manufacturing Engineering, National University of Sciences & Technology, Islamabad, Pakistan.

ABSTRACT
The COX-2/PGE₂ pathway has been implicated in the occurrence and progression of cancer. The underlying mechanisms facilitating the production of COX-2 and its mediator, PGE₂, in cancer survival remain unknown. Herein, we investigated PGE₂-induced COX-2 expression and signaling in HL-60 cells following menadione treatment. Treatment with PGE₂ activated anti-apoptotic proteins such as Bcl-2 and Bcl-xL while reducing pro-apoptotic proteins, thereby enhancing cell survival. PGE₂ not only induced COX-2 expression, but also prevented casapse-3, PARP, and lamin B cleavage. Silencing and inhibition of COX-2 with siRNA transfection or treatment with indomethacin led to a pronounced reduction of the extracellular levels of PGEv, and restored the menadione- induced cell death. In addition, pretreatment of cells with the MEK inhibitor PD98059 and the PKA inhibitor H89 abrogated the PGE₂-induced expression of COX-2, suggesting involvement of the MAPK and PKA pathways. These results demonstrate that PGE₂ signaling acts in an autocrine manner, and specific inhibition of PGE₂ will provide a novel approach for the treatment of leukemia.

Show MeSH
Related in: MedlinePlus