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Differential expression of apoptotic genes PDIA3 and MAP3K5 distinguishes between low- and high-risk prostate cancer.

Pressinotti NC, Klocker H, Schäfer G, Luu VD, Ruschhaupt M, Kuner R, Steiner E, Poustka A, Bartsch G, Sültmann H - Mol. Cancer (2009)

Bottom Line: These genes were analyzed by statistical, pathway and gene enrichment methods.Twenty selected candidate genes were verified by quantitative real time PCR and immunohistochemistry.In concordance with the mRNA levels, two genes MAP3K5 and PDIA3 exposed differential protein expression.

View Article: PubMed Central - HTML - PubMed

Affiliation: German Cancer Research Center, Division of Molecular Genome Analysis, Im Neuenheimer Feld 580, D-69120 Heidelberg, Germany. nicole@diergardt.eu

ABSTRACT

Background: Despite recent progress in the identification of genetic and molecular alterations in prostate cancer, markers associated with tumor progression are scarce. Therefore precise diagnosis of patients and prognosis of the disease remain difficult. This study investigated novel molecular markers discriminating between low and highly aggressive types of prostate cancer.

Results: Using 52 microdissected cell populations of low- and high-risk prostate tumors, we identified via global cDNA microarrays analysis almost 1200 genes being differentially expressed among these groups. These genes were analyzed by statistical, pathway and gene enrichment methods. Twenty selected candidate genes were verified by quantitative real time PCR and immunohistochemistry. In concordance with the mRNA levels, two genes MAP3K5 and PDIA3 exposed differential protein expression. Functional characterization of PDIA3 revealed a pro-apoptotic role of this gene in PC3 prostate cancer cells.

Conclusions: Our analyses provide deeper insights into the molecular changes occurring during prostate cancer progression. The genes MAP3K5 and PDIA3 are associated with malignant stages of prostate cancer and therefore provide novel potential biomarkers.

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siRNA mediated knockdown of PDIA3 decreased apoptosis in prostate cancer cell line. (A) Knockdown efficiency measured via qRT PCR 48 h after transfection with 20 nM siRNA. (B) PC3 cells were treated with 20 nM scrambled siRNA control and PDIA3 siRNA. 48 h after transfection induction of apoptosis was performed with 1 μM Staurosporine (STS), 20 μM Fenretinide (FenR) or 1.5 μM Tapsigargin (TG) for 6 and 24 hours. Apoptosis was measured by determining caspase activation and compared to untreated control. Bar heights and error bars are means and upper range of triplicate samples relative to control treatment. * P < 0.05 (unpaired t-test).
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Figure 4: siRNA mediated knockdown of PDIA3 decreased apoptosis in prostate cancer cell line. (A) Knockdown efficiency measured via qRT PCR 48 h after transfection with 20 nM siRNA. (B) PC3 cells were treated with 20 nM scrambled siRNA control and PDIA3 siRNA. 48 h after transfection induction of apoptosis was performed with 1 μM Staurosporine (STS), 20 μM Fenretinide (FenR) or 1.5 μM Tapsigargin (TG) for 6 and 24 hours. Apoptosis was measured by determining caspase activation and compared to untreated control. Bar heights and error bars are means and upper range of triplicate samples relative to control treatment. * P < 0.05 (unpaired t-test).

Mentions: MAP3K5 and PDIA3 were found to be proteins associated with apoptotic processes via pathway analysis. Unlike MAP3K5, whose pro-apoptotic and inflammatory role in context of tumorigenesis is well established [16,17], the function for PDIA3 in apoptosis has been largely unexplored. To investigate an apoptosis-related function of PDIA3 we performed siRNA-based knockdown in the human prostate cancer cell lines PC3 and LNCaP. 48 hours after siRNA treatment (20 nM or 40 nM) the knockdown efficiency was determined by qRT-PCR (Figure 4A). Induction of apoptosis was mediated by three different stimuli. Staurosporine (STS), Fenretinide (FenR) and Tapsigargin (TG) are known to activate apoptosis via distinct mechanisms [18-20]. Each stimulus activated the apoptotic pathway reflected by activation of caspase 3 and/or caspase 7 (Figure 4B). PDIA3 siRNA treatment revealed a significant decrease of caspase activation in PC3 cells with all stimuli in comparison to control siRNA treated cells. In LNCaP cells similar results were obtained, but were only significant after STS induction (data not shown). These results indicate a novel, pro-apoptotic role for PDIA3 in prostate cancer cells.


Differential expression of apoptotic genes PDIA3 and MAP3K5 distinguishes between low- and high-risk prostate cancer.

Pressinotti NC, Klocker H, Schäfer G, Luu VD, Ruschhaupt M, Kuner R, Steiner E, Poustka A, Bartsch G, Sültmann H - Mol. Cancer (2009)

siRNA mediated knockdown of PDIA3 decreased apoptosis in prostate cancer cell line. (A) Knockdown efficiency measured via qRT PCR 48 h after transfection with 20 nM siRNA. (B) PC3 cells were treated with 20 nM scrambled siRNA control and PDIA3 siRNA. 48 h after transfection induction of apoptosis was performed with 1 μM Staurosporine (STS), 20 μM Fenretinide (FenR) or 1.5 μM Tapsigargin (TG) for 6 and 24 hours. Apoptosis was measured by determining caspase activation and compared to untreated control. Bar heights and error bars are means and upper range of triplicate samples relative to control treatment. * P < 0.05 (unpaired t-test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2807430&req=5

Figure 4: siRNA mediated knockdown of PDIA3 decreased apoptosis in prostate cancer cell line. (A) Knockdown efficiency measured via qRT PCR 48 h after transfection with 20 nM siRNA. (B) PC3 cells were treated with 20 nM scrambled siRNA control and PDIA3 siRNA. 48 h after transfection induction of apoptosis was performed with 1 μM Staurosporine (STS), 20 μM Fenretinide (FenR) or 1.5 μM Tapsigargin (TG) for 6 and 24 hours. Apoptosis was measured by determining caspase activation and compared to untreated control. Bar heights and error bars are means and upper range of triplicate samples relative to control treatment. * P < 0.05 (unpaired t-test).
Mentions: MAP3K5 and PDIA3 were found to be proteins associated with apoptotic processes via pathway analysis. Unlike MAP3K5, whose pro-apoptotic and inflammatory role in context of tumorigenesis is well established [16,17], the function for PDIA3 in apoptosis has been largely unexplored. To investigate an apoptosis-related function of PDIA3 we performed siRNA-based knockdown in the human prostate cancer cell lines PC3 and LNCaP. 48 hours after siRNA treatment (20 nM or 40 nM) the knockdown efficiency was determined by qRT-PCR (Figure 4A). Induction of apoptosis was mediated by three different stimuli. Staurosporine (STS), Fenretinide (FenR) and Tapsigargin (TG) are known to activate apoptosis via distinct mechanisms [18-20]. Each stimulus activated the apoptotic pathway reflected by activation of caspase 3 and/or caspase 7 (Figure 4B). PDIA3 siRNA treatment revealed a significant decrease of caspase activation in PC3 cells with all stimuli in comparison to control siRNA treated cells. In LNCaP cells similar results were obtained, but were only significant after STS induction (data not shown). These results indicate a novel, pro-apoptotic role for PDIA3 in prostate cancer cells.

Bottom Line: These genes were analyzed by statistical, pathway and gene enrichment methods.Twenty selected candidate genes were verified by quantitative real time PCR and immunohistochemistry.In concordance with the mRNA levels, two genes MAP3K5 and PDIA3 exposed differential protein expression.

View Article: PubMed Central - HTML - PubMed

Affiliation: German Cancer Research Center, Division of Molecular Genome Analysis, Im Neuenheimer Feld 580, D-69120 Heidelberg, Germany. nicole@diergardt.eu

ABSTRACT

Background: Despite recent progress in the identification of genetic and molecular alterations in prostate cancer, markers associated with tumor progression are scarce. Therefore precise diagnosis of patients and prognosis of the disease remain difficult. This study investigated novel molecular markers discriminating between low and highly aggressive types of prostate cancer.

Results: Using 52 microdissected cell populations of low- and high-risk prostate tumors, we identified via global cDNA microarrays analysis almost 1200 genes being differentially expressed among these groups. These genes were analyzed by statistical, pathway and gene enrichment methods. Twenty selected candidate genes were verified by quantitative real time PCR and immunohistochemistry. In concordance with the mRNA levels, two genes MAP3K5 and PDIA3 exposed differential protein expression. Functional characterization of PDIA3 revealed a pro-apoptotic role of this gene in PC3 prostate cancer cells.

Conclusions: Our analyses provide deeper insights into the molecular changes occurring during prostate cancer progression. The genes MAP3K5 and PDIA3 are associated with malignant stages of prostate cancer and therefore provide novel potential biomarkers.

Show MeSH
Related in: MedlinePlus