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Farnesoid X receptor, overexpressed in pancreatic cancer with lymph node metastasis promotes cell migration and invasion.

Lee JY, Lee KT, Lee JK, Lee KH, Jang KT, Heo JS, Choi SH, Kim Y, Rhee JC - Br. J. Cancer (2011)

Bottom Line: The microarray results were validated by real-time PCR.Small interfering RNA-mediated downregulation of FXR and guggulsterone-mediated FXR inhibition resulted in a marked reduction in cell migration and invasion.In addition, downregulation of FXR reduced NF-κB activation and conditioned medium from FXR siRNA-transfected cells showed reduced VEGF levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Health Science and Technology, Graduate School, SAIHST, Sungkyunkwan University, Seoul, Korea.

ABSTRACT

Background: Lymph node metastasis is one of the most important adverse prognostic factors for pancreatic cancer. The aim of this study was to identify novel lymphatic metastasis-associated markers and therapeutic targets for pancreatic cancer.

Methods: DNA microarray study was carried out to identify genes differentially expressed between 17 pancreatic cancer tissues with lymph node metastasis and 17 pancreatic cancer tissues without lymph node metastasis. The microarray results were validated by real-time PCR. Immunohistochemistry and western blotting were used to examine the expression of farnesoid X receptor (FXR). The function of FXR was studied by small interfering RNA and treatment with FXR antagonist guggulsterone and FXR agonist GW4064.

Results: Farnesoid X receptor overexpression in pancreatic cancer tissues with lymph node metastasis is associated with poor patient survival. Small interfering RNA-mediated downregulation of FXR and guggulsterone-mediated FXR inhibition resulted in a marked reduction in cell migration and invasion. In addition, downregulation of FXR reduced NF-κB activation and conditioned medium from FXR siRNA-transfected cells showed reduced VEGF levels. Moreover, GW4064-mediated FXR activation increased cell migration and invasion.

Conclusions: These findings indicated that FXR overexpression plays an important role in lymphatic metastasis of pancreatic cancer and that downregulation of FXR is an effective approach for inhibition of pancreatic tumour progression.

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Related in: MedlinePlus

Gene expression analysis of selected upregulated genes and immunohistochemical expression of FXR in pancreatic cancer tissues with and without LN metastasis. (A) Upregulated gene analysis between pancreatic cancer tissues with and without LN metastasis by real-time PCR. (B) Real-time PCR validation of microarray data. For each sample, the amount of target and endogenous reference was determined from the appropriate standard curve. The target amount was then divided by an endogenous reference (GAPDH) amount to obtain a normalised target value. The black columns represent the mean of fold (Group II/Group I) values obtained in DNA microarray; the grey columns represent the mean of fold (Group II/Group I) values obtained in real-time PCR; *P<0.05. (C) Overexpression of FXR in pancreatic cancer tissues with LN metastasis. The FXR intensity on tissues was evaluated by immunohistochemical staining. (a) A representative negative staining in pancreatic cancer tissues without LN metastasis. Magnification, × 200. (b) A representative positive staining in pancreatic cancer tissues with LN metastasis. Magnification, × 200. (D) Kaplan–Meier analysis of overall survival for patients with ductal adenocarcinoma (n=34). Farnesoid X receptor expression-positive cases (n=15) had significantly poorer prognosis than expression -negative cases (n=19; P=0.0227, log-rank test).
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fig1: Gene expression analysis of selected upregulated genes and immunohistochemical expression of FXR in pancreatic cancer tissues with and without LN metastasis. (A) Upregulated gene analysis between pancreatic cancer tissues with and without LN metastasis by real-time PCR. (B) Real-time PCR validation of microarray data. For each sample, the amount of target and endogenous reference was determined from the appropriate standard curve. The target amount was then divided by an endogenous reference (GAPDH) amount to obtain a normalised target value. The black columns represent the mean of fold (Group II/Group I) values obtained in DNA microarray; the grey columns represent the mean of fold (Group II/Group I) values obtained in real-time PCR; *P<0.05. (C) Overexpression of FXR in pancreatic cancer tissues with LN metastasis. The FXR intensity on tissues was evaluated by immunohistochemical staining. (a) A representative negative staining in pancreatic cancer tissues without LN metastasis. Magnification, × 200. (b) A representative positive staining in pancreatic cancer tissues with LN metastasis. Magnification, × 200. (D) Kaplan–Meier analysis of overall survival for patients with ductal adenocarcinoma (n=34). Farnesoid X receptor expression-positive cases (n=15) had significantly poorer prognosis than expression -negative cases (n=19; P=0.0227, log-rank test).

Mentions: The differences in gene expression found by microarray analyses were validated by real-time PCR. We chose genes that were >2-fold upregulated according to our microarray results (Table 2). Among the genes, we evaluated the levels of expression of four selected upregulated genes in pancreatic cancer tissues with LN metastasis. The expression of LZIC, FXR, SCAMP1, and SULT1E1 was significantly higher in pancreatic cancer tissues with LN metastasis than in pancreatic cancer tissues without LN metastasis (Figure 1A). Although some variation concerning the degree of regulation was observed, the data obtained with microarrays were substantially confirmed for four selective genes by real-time PCR (Figure 1B). In summary, microarrays together with real-time PCR validation results clearly show that the expression of LZIC, FXR, SCAMP1, and SULT1E1 were confirmed to be significantly higher in pancreatic cancer tissues with LN metastasis than in pancreatic cancer tissues without LN metastasis. These results are representative for measurements accomplished with pancreatic cancer tissues from three different experiments.


Farnesoid X receptor, overexpressed in pancreatic cancer with lymph node metastasis promotes cell migration and invasion.

Lee JY, Lee KT, Lee JK, Lee KH, Jang KT, Heo JS, Choi SH, Kim Y, Rhee JC - Br. J. Cancer (2011)

Gene expression analysis of selected upregulated genes and immunohistochemical expression of FXR in pancreatic cancer tissues with and without LN metastasis. (A) Upregulated gene analysis between pancreatic cancer tissues with and without LN metastasis by real-time PCR. (B) Real-time PCR validation of microarray data. For each sample, the amount of target and endogenous reference was determined from the appropriate standard curve. The target amount was then divided by an endogenous reference (GAPDH) amount to obtain a normalised target value. The black columns represent the mean of fold (Group II/Group I) values obtained in DNA microarray; the grey columns represent the mean of fold (Group II/Group I) values obtained in real-time PCR; *P<0.05. (C) Overexpression of FXR in pancreatic cancer tissues with LN metastasis. The FXR intensity on tissues was evaluated by immunohistochemical staining. (a) A representative negative staining in pancreatic cancer tissues without LN metastasis. Magnification, × 200. (b) A representative positive staining in pancreatic cancer tissues with LN metastasis. Magnification, × 200. (D) Kaplan–Meier analysis of overall survival for patients with ductal adenocarcinoma (n=34). Farnesoid X receptor expression-positive cases (n=15) had significantly poorer prognosis than expression -negative cases (n=19; P=0.0227, log-rank test).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3065277&req=5

fig1: Gene expression analysis of selected upregulated genes and immunohistochemical expression of FXR in pancreatic cancer tissues with and without LN metastasis. (A) Upregulated gene analysis between pancreatic cancer tissues with and without LN metastasis by real-time PCR. (B) Real-time PCR validation of microarray data. For each sample, the amount of target and endogenous reference was determined from the appropriate standard curve. The target amount was then divided by an endogenous reference (GAPDH) amount to obtain a normalised target value. The black columns represent the mean of fold (Group II/Group I) values obtained in DNA microarray; the grey columns represent the mean of fold (Group II/Group I) values obtained in real-time PCR; *P<0.05. (C) Overexpression of FXR in pancreatic cancer tissues with LN metastasis. The FXR intensity on tissues was evaluated by immunohistochemical staining. (a) A representative negative staining in pancreatic cancer tissues without LN metastasis. Magnification, × 200. (b) A representative positive staining in pancreatic cancer tissues with LN metastasis. Magnification, × 200. (D) Kaplan–Meier analysis of overall survival for patients with ductal adenocarcinoma (n=34). Farnesoid X receptor expression-positive cases (n=15) had significantly poorer prognosis than expression -negative cases (n=19; P=0.0227, log-rank test).
Mentions: The differences in gene expression found by microarray analyses were validated by real-time PCR. We chose genes that were >2-fold upregulated according to our microarray results (Table 2). Among the genes, we evaluated the levels of expression of four selected upregulated genes in pancreatic cancer tissues with LN metastasis. The expression of LZIC, FXR, SCAMP1, and SULT1E1 was significantly higher in pancreatic cancer tissues with LN metastasis than in pancreatic cancer tissues without LN metastasis (Figure 1A). Although some variation concerning the degree of regulation was observed, the data obtained with microarrays were substantially confirmed for four selective genes by real-time PCR (Figure 1B). In summary, microarrays together with real-time PCR validation results clearly show that the expression of LZIC, FXR, SCAMP1, and SULT1E1 were confirmed to be significantly higher in pancreatic cancer tissues with LN metastasis than in pancreatic cancer tissues without LN metastasis. These results are representative for measurements accomplished with pancreatic cancer tissues from three different experiments.

Bottom Line: The microarray results were validated by real-time PCR.Small interfering RNA-mediated downregulation of FXR and guggulsterone-mediated FXR inhibition resulted in a marked reduction in cell migration and invasion.In addition, downregulation of FXR reduced NF-κB activation and conditioned medium from FXR siRNA-transfected cells showed reduced VEGF levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Health Science and Technology, Graduate School, SAIHST, Sungkyunkwan University, Seoul, Korea.

ABSTRACT

Background: Lymph node metastasis is one of the most important adverse prognostic factors for pancreatic cancer. The aim of this study was to identify novel lymphatic metastasis-associated markers and therapeutic targets for pancreatic cancer.

Methods: DNA microarray study was carried out to identify genes differentially expressed between 17 pancreatic cancer tissues with lymph node metastasis and 17 pancreatic cancer tissues without lymph node metastasis. The microarray results were validated by real-time PCR. Immunohistochemistry and western blotting were used to examine the expression of farnesoid X receptor (FXR). The function of FXR was studied by small interfering RNA and treatment with FXR antagonist guggulsterone and FXR agonist GW4064.

Results: Farnesoid X receptor overexpression in pancreatic cancer tissues with lymph node metastasis is associated with poor patient survival. Small interfering RNA-mediated downregulation of FXR and guggulsterone-mediated FXR inhibition resulted in a marked reduction in cell migration and invasion. In addition, downregulation of FXR reduced NF-κB activation and conditioned medium from FXR siRNA-transfected cells showed reduced VEGF levels. Moreover, GW4064-mediated FXR activation increased cell migration and invasion.

Conclusions: These findings indicated that FXR overexpression plays an important role in lymphatic metastasis of pancreatic cancer and that downregulation of FXR is an effective approach for inhibition of pancreatic tumour progression.

Show MeSH
Related in: MedlinePlus