Effects of parthenolide on the expression of pro-inflammatory genes.

<p>LNCaP (<b>A</b>), PC3 (<b>B</b>), DU145 (<b>C</b>) cells were kept in normoxia and exposed to 1% O₂ in the presence and absence of 5 µM parthenolide. mRNA levels for VEGF, RAGE, P2X7R, PTX3, COX2, CXCR4 and HO1 were analyzed by real-time PCR, normalized to the housekeeping gene 18S rRNA and expressed as fold induction with respect to the normoxic untreated control (set at 1). For each gene we present the effect of parthenolide at the time points where hypoxia exerted a maximum increase on its transcription. *Hypoxic parthenolide treated cells <i>vs</i> hypoxic cells. C: control normoxic untreated cells. P: normoxic parthenolide treated cells. H: hypoxic cells. HP: parthenolide treated hypoxic cells.</p

Effects of parthenolide on the expression of the anti-inflammatory gene HO1.

<p>LNCaP, PC3 and DU145 cells were kept in normoxia and exposed to 1% O₂ in the presence and absence of 5 µM parthenolide. mRNA levels for HO1 were analyzed by real-time PCR, normalized to the housekeeping gene 18S rRNA and expressed as fold induction with respect to the normoxic untreated control (set at 1). We show the effect of parthenolide after 4 h treatment when the drug maximally upregulated HO1 expression in normoxic cultures and after 24 h (LNCaP, DU145) and 48 h (PC3) stimulation when hypoxia exerted a maximum increase on its transcription. Mean values ± SE. * Normoxic and hypoxic parthenolide treated cells <i>vs</i> normoxic control cells. C: control normoxic untreated cells. P: normoxic parthenolide treated cells. H: hypoxic cells. HP: parthenolide treated hypoxic cells.</p

Hypoxic regulation of NF-kB nuclear expression in LNCaP, DU145 and PC3 cells.

<p>Cell were exposed to 1% O₂ from 0.5 up to 24 h or left untreated. After the times indicated, cells were processed and the nuclear content of p50 and p65 was detected by immunoblot analysis. β-actin was used as a loading control. A representative experiment for each gene in all the studied cell lines is shown. Mean densitometry of NF-kB p50 and p65 relative to β-actin ± SE is also evidenced and expressed as arbitrary units. * Hypoxic cells <i>vs</i> normoxic control cells.</p

Effects of Parthenolide on HIF1α and HIF3α hypoxia dependent activation.

<p>A LNCaP, DU145 and PC3 cells were kept in normoxia and exposed to 1% O₂ for 4 h in the presence and absence of 5 µM parthenolide. HIF1α content was measured in the nuclear fraction by immunoblot analysis. β-actin was used as a loading control. For each gene a representative experiment is shown. Mean densitometry of HIF1α relative to β-actin is also evidenced and expressed as arbitrary units. <b>B</b> DU145 cells were incubated under normoxic or hypoxic conditions for 48 h in the presence and absence of 5 µM parthenolide. HIF3α content was measured in the nuclear fraction by immunoblot analysis. β-actin was used as a loading control. For each gene a representative experiment is shown. Mean densitometry of HIF3α relative to β-actin is also evidenced and expressed as arbitrary units. Mean values ± SE. C: control normoxic untreated cells. P: normoxic parthenolide treated cells. H: hypoxic cells. HP: parthenolide treated hypoxic cells. *Hypoxic parthenolide treated cells <i>vs</i> hypoxic cells.</p

mRNA expression levels of pro-inflammatory genes in hypoxic LNCaP, DU145 and PC3 cells.

<p>Cells were exposed to 1% O₂ from 2 up to 72 h. mRNA levels for VEGF, RAGE, P2X7R, PTX3, COX2, CXCR4 and HO1 were analyzed by real-time PCR and normalized to the housekeeping gene 18S rRNA. Graphs show the ratio between the expression of hypoxia treated cells with respect to normoxic control cells (set at 1). Mean values ± SE. * Hypoxic cells <i>vs</i> normoxic control cells.</p

Effects of HIF1α knockdown and of parthenolide on the normoxic and hypoxic expression of the pro-inflammatory phenotype and HO1 transcription in DU145 cells.

<p><b>A</b> HIF1α shRNA cells were exposed to 1% O₂ or left untreated from 2 up to 72 h in the presence and absence of 5 µM parthenolide. mRNA levels for VEGF, RAGE, PTX3, COX2 and CXCR4 were analyzed by real-time PCR and normalized to the housekeeping gene 18S rRNA. For each gene we present the result on the time point where hypoxia exerted a maximum increase in transcription. Graphs show the ratio between the expression of parthenolide, hypoxia, parthenolide and hypoxia treated HIF1α shRNA cells with respect to the mean normoxic control of wt cell cultures (set at 1). Mean values ± SE. ° HIF1α shRNA normoxic control cells <i>vs</i> wt normoxic control cells, * hypoxic HIF1α shRNA cells <i>vs</i> HIF1α shRNA normoxic control cells. # Parthenolide treated hypoxic HIF1α shRNA cells <i>vs</i> hypoxic HIF1α shRNA cells. <b>B</b> Effect of parthenolide on the transcription the anti-inflammatory gene HO1 in HIF1α shRNA cells in normoxia and in hypoxia after 4 h (when parthenolide maximally upregulated HO1 expression in normoxic cultures) and 24 h treatment. Graph shows the ratio between the expression of parthenolide, hypoxia, parthenolide and hypoxia treated HIF1α shRNA cells with respect to the mean normoxic control of wt cell cultures (set at 1). Mean values ± SE. * Normoxic and hypoxic parthenolide treated HIF1α shRNA cells <i>vs</i> HIF1α shRNA normoxic control cells. C: control normoxic untreated cells. P: parthenolide treated cells. H: hypoxic cells. HP: parthenolide treated hypoxic cells.</p

Normoxic expression levels of pro-inflammatory genes in LNCaP, DU145 and PC3 cells.

<p>Normoxic expression levels of pro-inflammatory genes in LNCaP, DU145 and PC3 cells. Values express the mRNA level of each gene quantified by real-time PCR and normalized to the housekeeping gene 18S rRNA. Mean ± SE. Asterisks indicate a significant higher value compared to LNCaP cells.</p

Molecular analysis of the effects of Piroxicam and Cisplatin on mesothelioma cells growth and viability-2

Hase distribution. All the experiments were performed at least 3 times. Media ± Standard deviation of experiments is expressed as percentage of total cells. CTRL = control; P = piroxicam; C = CDDP.<p><b>Copyright information:</b></p><p>Taken from "Molecular analysis of the effects of Piroxicam and Cisplatin on mesothelioma cells growth and viability"</p><p>Journal of Translational Medicine 2008;6():27-27.</p><p>Published online 22 May 2008</p><p>PMCID:PMC2412853.</p><p></p

Molecular analysis of the effects of Piroxicam and Cisplatin on mesothelioma cells growth and viability-6

Ed by cell counts and was expressed as percent of control (untreated cells). Experiments were repeated in triplicate and media values were calculated and indicated in the upper table. P values at the different points of the treatments respect to the control and of the combined treatment (P+C) respect to the single drugs treatment were indicated in the lower tables. CTRL = control; P = piroxicam; C = CDDP.<p><b>Copyright information:</b></p><p>Taken from "Molecular analysis of the effects of Piroxicam and Cisplatin on mesothelioma cells growth and viability"</p><p>Journal of Translational Medicine 2008;6():27-27.</p><p>Published online 22 May 2008</p><p>PMCID:PMC2412853.</p><p></p

Molecular analysis of the effects of Piroxicam and Cisplatin on mesothelioma cells growth and viability-0

Ed by cell counts and was expressed as percent of control (untreated cells). Experiments were repeated in triplicate and media values were calculated and indicated in the upper table. P values at the different points of the treatments respect to the control and of the combined treatment (P+C) respect to the single drugs treatment were indicated in the lower tables. CTRL = control; P = piroxicam; C = CDDP.<p><b>Copyright information:</b></p><p>Taken from "Molecular analysis of the effects of Piroxicam and Cisplatin on mesothelioma cells growth and viability"</p><p>Journal of Translational Medicine 2008;6():27-27.</p><p>Published online 22 May 2008</p><p>PMCID:PMC2412853.</p><p></p