Apoptosis in PTJ64i and PTJ86i cells treated with 24 μM GW6471 for 24, 48 or 72 hours.

<p>Values represented in the histograms (<i>top</i>) are the means ±SD of at least two independent determinations (*<i>p</i><0.05; **<i>p</i><0.01; ***<i>p</i><0.001). Dot plots (<i>bottom</i>) show representative experiments after a 72-hour treatment with 24 μM GW6471.</p

Immunohistochemical analysis of PPARα in HNPGL tissues.

<p>Tumor sections derived from two different HNPGL patients show positive immunostaining for PPARα in paraganglioma cell clusters, while the supporting stroma is negative. Bar = 80 μm.</p

Immunofluorescence analysis of PPARs in HNPGL cells.

<p>PTJ64i (<i>left panels</i>) and PTJ86i (<i>right panels</i>) show high levels of PPARα nuclear expression. The fluorescence intensity for PPARα is stronger as compared to that for PPARβ/δ or PPARγ. Bar = 20 μm.</p

Growth curves of PTJ64i and PTJ86i cells treated with GW6471.

<p>Cell number was measured over a 72-hour time course treatment with 24 μM GW6471 or with vehicle control. Data shown are the means ±SD of three to five determinations (*<i>p</i><0.05; **<i>p</i><0.01; ***<i>p</i><0.001).</p

Effect of GW6471 on PI3K/GSK-3β/β-catenin signaling pathway in PTJ64i and PTJ86i cell lines.

<p>Histograms of normalized densitometric analyses and representative western blotting for PI3K, GSK3β, p-GSK3β and β-catenin in HNPGL cells treated with 24 μM GW6471 for 72 hours are shown. The relative densities of the immunoreactive bands were determined and normalized with respect to GAPDH (loading control) using a semiquantitative densitometric analysis (Kodak ID Image Analysis Software, Rochester, NY, USA). Values are expressed as relative units (RU). Each bar represents the mean ± SD of two up to five independent determinations (*<i>p</i><0.05).</p

Effect of GW6471 on clonogenic activity of PTJ64i and PTJ86i cell lines.

<p>Data shown in the histograms are the means ±SD of three independent experiments (*<i>p</i><0.05; **<i>p</i><0.01). PE: plating efficiency [(# of colonies formed/# of cells plated)*100]; SF: surviving fraction [# of colonies formed *100/(# of cells plated *PE of control vehicle)].</p

Effect of GW6471 on PTJ64i and PTJ86i cell cycles.

<p>The histograms show the mean percentage of cells (values inside the bars) in the different cell phases after a 24-hour treatment with 24 μM GW6471 as compared to control (**<i>p</i><0.01; ***<i>p</i><0.001).</p