Cancer Res Treat.  2004 Oct;36(5):330-337.

Dexamethasone Inhibits TRAIL- and Anti-cancer Drugs-induced Cell Death in A549 Cells through Inducing NF-kappaB-independent cIAP2 Expression

• Kim YS
• Park JS
• Jee YK
• Lee KY

Abstract

PURPOSE
We have examined that dexamethasone inhibits apoptotic cell death of A549 lung epithelial cells through TRAIL and anti-cancer drugs. The purpose of the study is to determine the roles of GR, cIAP and NF- kappaB in this mechanism. MATERIALS AND METHODS: In the A549 lung epithelial cell line, TRAIL, taxol, doxorubicine & gemcitabine were used to investigate cell toxicity. Cells were pretreated 12 hours in advance with dexamethasone. RU486 was pretreated 30 minutes before dexamethasone. Crystal violet assay was used for cell toxicity tests. Apoptosis assay was performed by taking morphologic surveys with fluorescent microscopy after double staining with Hoechst 33342 & propium iodide. RT-PCR was used to investigate the gene expression of cIAP1 & cIAP2 by dexamethasone. Ad-IkappaBalpha-SR transduction study was used for the role of NF-kappaB. RESULTS: TRAIL and anti-cancer drug-induced apoptosis was partially suppressed in A549 cells pretreated with dexamethasone. The inhibitory effect on cell death disappeared in A549 cells pretreated with RU486. Using RT-PCR, changes of cIAP1 and cIAP2 genes manifestation in A549 cells subsequent to pretreatment with dexamethasone were examined. The results showed an increase in cIAP2 expression during a course of time which was suppressed by RU486 pretreatment. Induction of cIAP2 expression changes by dexamethasone was uniquely observed despite the blockade of NF-kappa by Ad-IkappaB alpha-SR transduction. CONCLUSIONS: These results suggest that dexametha sone inhibits TRAIL- and anti-cancer drug-induced apoptosis in A549 cells by inducing cIAP2 gene expression through a GR-mediated, NF-kappa-independent pathway.

Keyword

Apoptotis; A549 cells; cIAP; Dexamethasone; GR; NF-kappaB

MeSH Terms

Apoptosis
Cell Death*
Dexamethasone*
Doxorubicin
Epithelial Cells
Gene Expression
Gentian Violet
Lung
Microscopy
Mifepristone
NF-kappa B
Paclitaxel
Prednisone
Toxicity Tests
Dexamethasone
Doxorubicin
Gentian Violet
Mifepristone
NF-kappa B
Paclitaxel
Prednisone

Figure

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  Fig. 1 Dexamethasone inhibits TRAIL-induced cell death in A549 cells. A549 cells were treated with control medium, dexamethasone alone, TRAIL alone, and dexamethasone for 12 hr before the addition of TRAIL. Cell viability was assessed by light microscopy 24 hr later.

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  Fig. 2 Dexamethasone inhibits anti-cancer drug-induced cell death in A549 cells. A549 cells were treated with control medium, dexamethasone and/or anti-cancer drugs: taxol, doxorubicine, and gemcitabine for 24 hr followed by analysis of cell viability by crystal violet assay. In experiments with dexamethasone plus anti-cancer drugs, A549 cells were pretreated with dexamethasone for 12 hr before the addition of anti-cancer drugs, simultaneously treated with dexamethasone and anti-cancer drugs, and posttreated with dexamethasone for 12 hr after the addition of anti-cancer drugs.

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  Fig. 3 Induction of cIAP2 mRNA expression by dexamethasone in A549 cells. A549 cells were treated with dexamethasone and harvested at 0, 1 hr, 2 hr, 4 hr for RT-PCR analysis with a cIAP1 primer, a cIAP2 primer, and a beta-actin primer.

• 

  Fig. 4 RU486 blocks inhibition of TRAIL-induced cell death by dexamethasone in A549 cells. A549 cells were treated with control medium, TRAIL, pretreated with dexamethasone 12 hr before the addition of TRAIL and pretreated with RU486 30 min before the addition of dexamethasone and TRAIL. Cell viability was assessed by crystal violet assay 24 hr later.

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  Fig. 5 RU486 inhibits dexamethasone-induced cIAP2 mRNA expression in A549 cells. A549 cells were treated with dexamethasone, RU486 plus dexamethasone, and harvested at 0, 1 hr, 2 hr, 4 hr for RT-PCR analysis with a cIAP2 primer and beta-actin primer. In experiments with RU486 plus dexamethasone, A549 cells were pretreated with RU486 for 30 min before the addition of dexamethasone.

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  Fig. 6 NF-κB has no effect on inhibition of TRAIL-induced cell death by dexamethasone in A549 cells. A549 cells were infected with Ad-null or Ad-IκBα-SR at 20 m.o.i for 20 hr and then incubated with TRAIL or 12 hr pretreatment with dexamethasone. Cell viability was assessed by crystal violet assay 24 hr later.

• 

  Fig. 7 No effect of NF-κB on dexamethasone-induced cIAP2 mRNA expression in A549 cells. A549 cells were infected with Ad-null or Ad-IκBα-SR at 20 m.o.i for 20 hr, then incubated with dexamethasone and harvested at 0, 1 hr, 2 hr, 4 hr for RT-PCR analysis with cIAP2 primer and beta-actin primer.

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