Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: Sequences of primers.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Sequencing, Amplification

Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: HPA and FGF2 are upregulated in pancreatic cancer tissues and cell lines. (A) Comparison of HPA and FGF2 expression in pancreatic cancer tissues and adjacent normal pancreatic tissues by immunohistochemical stainin. (B) Analysis of relative mRNA expression levels of HPA and FGF2 in pancreatic cancer and normal pancreatic cell lines by reverse transcription-quantitative PCR. *P<0.05, **P<0.01 vs. HPDE6c7. HPA, heparanase; FGF2, fibroblast growth factor 2; NC, negative control.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Comparison, Expressing, Immunohistochemical staining, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control

Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: Expression of heparanase and fibroblast growth factor 2 in pancreatic cancer tissues and adjacent normal tissues.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Expressing

Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: Association between heparanase and fibroblast growth factor 2 expression, and clinicopathological characteristics of patients with pancreatic cancer.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Expressing

Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: HPA upregulates the expression of FGF2 in pancreatic cancer cell lines. (A) Analysis of HPA silencing and overexpression efficiency by RT-qPCR. (B) Analysis of HPA silencing and overexpression efficiency by western blot assay. (C) Analysis of FGF2 expression after silencing or overexpressing HPA by RT-qPCR. (D) Analysis of FGF2 expression after silencing or overexpressing HPA by western blotting. β-actin was used as an internal control. *P<0.05, **P<0.01, ***P<0.001. RT-qPCR, reverse transcription-quantitative PCR; HPA, heparanase; FGF2, fibroblast growth factor 2; sh, short hairpin (RNA; specific for HPA); Vector, PANC-1 cell line transfected with empty plasmid containing the GFP sequence.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Expressing, Over Expression, Quantitative RT-PCR, Western Blot, Control, Reverse Transcription, Real-time Polymerase Chain Reaction, shRNA, Plasmid Preparation, Transfection, Sequencing

Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: HPA regulates FGF2 expression via the HPA/SDC1 axis in pancreatic cancer cell lines. (A) RT-qPCR comparing the relative expression levels of SDC1 mRNA in pancreatic cancer lines and the normal pancreatic cell line HPDE6c7. *P<0.05, **P<0.01 vs. HPDE6c7. (B) Detection of the effect of HPA on SDC1 mRNA expression by RT-qPCR. (C) Expression of FGF2 was determined by RT-qPCR analysis after adding the SDC1 inhibitor synstatin to the HPA overexpression group. **P<0.01. RT-qPCR, reverse transcription-quantitative PCR; HPA, heparanase; FGF2, fibroblast growth factor 2; sh, short hairpin (RNA); SDC1, syndecan-1; Vector, PANC-1 cell line transfected with empty plasmid containing the GFP sequence.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Expressing, Quantitative RT-PCR, Over Expression, Reverse Transcription, Real-time Polymerase Chain Reaction, shRNA, Plasmid Preparation, Transfection, Sequencing

Journal: Oncology Letters

Article Title: The HPA/SDC1 axis promotes invasion and metastasis of pancreatic cancer cells by activating EMT via FGF2 upregulation

doi: 10.3892/ol.2019.11121

Figure Lengend Snippet: HPA promotes cell migration and invasion by activating the PI3K/Akt signaling pathway and EMT processes via FGF2 upregulation. (A) Expression of Palladin protein in various pancreatic cancer cell lines was analyzed by western blot assay. β-actin was used as an internal control. (B) Analysis of the relative expression of Palladin mRNA in pancreatic cancer cell lines by reverse transcription-quantitative PCR. (C) Western blot analysis of the effect of FGF2 on Palladin and Akt. (D) Protein expression of Palladin and Akt. β-actin was used as an internal control. (E) Effect of FGF2, AZD4547 and LY294002 on the migratory ability of PANC-1 cells assessed by wound healing assay. Magnification, ×200. (F) Transwell assays were performed to determine the invasive ability of PANC-1 cells treated with FGF2, AZD4547 and LY294002. (G) Number of invaded PANC-1 cells treated with FGF2, AZD4547 and LY294002. CON, PANC-1 cell line transfected with empty plasmid. (H) Western blot analysis of the effect of HPA and FGF2 on EMT. Vector, PANC-1 cell line transfected with empty plasmid. β-actin was used as an internal control. *P<0.05, **P<0.01, ***P<0.001. HPA, heparanase; FGF2, fibroblast growth factor 2; EMT, epithelial-mesenchymal transition; CON, PANC-1 cell line transfected with empty plasmid; Vector, PANC-1 cell line transfected with empty plasmid containing the GFP sequence.

Article Snippet: After blocking with 5% skimmed milk for 2 h at room temperature, the membranes were incubated with the following primary antibodies: Rabbit anti-human HPA (1:1,000; cat. no. PB0405), rabbit anti-human FGF2 (1:1,000; cat. no. PB0916), rabbit anti-human AKT (1:5,000; cat. no. A00024-1), rabbit anti-human E-cadherin (1:100; cat. no. BA0475), rabbit anti-human N-cadherin (1:100; cat. no. BM3921), rabbit anti-human vimentin 1:3,000 (cat. no. PB0378; all Boster Biological Technology) and rabbit anti-human palladin (1:100; cat. no. PA5-65160; Thermo Fisher Scientific Inc.) were added according to the instructions of the manufacturer and incubated at 4°C overnight. β-actin (1:500; cat. no. BA2305; Boster Biological Technology) was used as an internal loading control.

Techniques: Migration, Expressing, Western Blot, Control, Reverse Transcription, Real-time Polymerase Chain Reaction, Wound Healing Assay, Transfection, Plasmid Preparation, Sequencing