J Korean Med Sci.  2016 Oct;31(10):1586-1594. 10.3346/jkms.2016.31.10.1586.

Eukaryotic Translation Initiation Factor 3a (eIF3a) Promotes Cell Proliferation and Motility in Pancreatic Cancer

• Wang SQ
• Liu Y
• Yao MY
• Jin J

• KMID: 2468249
• DOI: http://doi.org/10.3346/jkms.2016.31.10.1586

Abstract

Identifying a target molecule that is crucially involved in pancreatic tumor growth and metastasis is necessary in developing an effective treatment. The study aimed to investigate the role of the eukaryotic translation initiation factor 3a (eIF3a) in the cell proliferation and motility in pancreatic cancer. Our data showed that the expression of eIF3a was upregulated in pancreatic ductal adenocarcinoma as compared with its expression in normal pancreatic tissues. Knockdown of eIF3a by a specific shRNA caused significant decreases in cell proliferation and clonogenic abilities in pancreatic cancer SW1990 and Capan-1 cells. Consistently, the pancreatic cancer cell growth rates were also impaired in xenotransplanted mice. Moreover, wound-healing assay showed that depletion of eIF3a significantly slowed down the wound recovery processes in SW1990 and Capan-1 cells. Transwell migration and invasion assays further showed that cell migration and invasion abilities were significantly inhibited by knockdown of eIF3a in SW1990 and Capan-1 cells. Statistical analysis of eIF3a expression in 140 cases of pancreatic ductal adenocarcinoma samples revealed that eIF3a expression was significantly associated with tumor metastasis and TNM staging. These analyses suggest that eIF3a contributes to cell proliferation and motility in pancreatic ductal adenocarcinoma.

Keyword

eIF3a; Proliferation; Migration; Invasion; Pancreatic Ductal Adenocarcinoma

MeSH Terms

Aged
Animals
Carcinoma, Pancreatic Ductal/metabolism/*pathology/therapy
Cell Line, Tumor
Cell Movement
Cell Proliferation
Eukaryotic Initiation Factor-3/antagonists & inhibitors/genetics/*metabolism
Female
Humans
Immunohistochemistry
Male
Mice
Mice, Inbred BALB C
Mice, Nude
Middle Aged
Pancreatic Neoplasms/metabolism/*pathology/therapy
RNA Interference
RNA, Small Interfering/administration & dosage/metabolism
Transplantation, Heterologous
Eukaryotic Initiation Factor-3
RNA, Small Interfering

Figure

• 

  Fig. 1 Aberrant eIF3a expression in pancreatic cancer tissues. (A) Relative eIF3a mRNA levels in the pancreatic ductal adenocarcinoma tissues (n = 30) and their paired adjacent non-cancerous pancreas tissues (n = 30). (B) Immunohistochemistry analysis of the protein level of eIF3a in slides from normal pancreas tissues and the pancreatic ductal adenocarcinoma tissues (n = 140). It was shown that eIF3a was non-expressed or lowly expressed in normal pancreatic duct (black arrow) and normal pancreatic acini (red arrow). In contrast, eIF3a was strongly expressed in the cancer tissues. *P < 0.001

• 

  Fig. 2 The constitutive expression of eIF3a in pancreatic cancer cell lines and the knockdown efficacy of a specific shRNA against eIF3a. (A) In the seven pancreatic cancer cell lines, it was observed that SW1990 and Capan-1 cells exhibited the strongest expression of eIF3a, whereas Miapaca-2 cell line exhibited the least expression of eIF3a. Hence, SW1990 and Capan-1 were chosen for subsequent analyses. (B) A specific shRNA against eIF3a was utilized to knock down the expression of eIF3a in SW1990 cells and Capan-1 cells. Western blot analysis revealed that the protein level of eIF3a was barely detected after transfection of the specific shRNA into these two cell lines, suggesting the high efficacy of our designed shRNA.

• 

  Fig. 3 Knockdown of eIF3a inhibited cell proliferation and colony formation in pancreatic cancer cell lines. (A) In the cell proliferation assay, significant disparities were observed from the fourth day in SW1990 cells and the fifth day in Capan-1 cells. The inhibitory effects grew as the time extended in both cells. (B) In the colony formation assay, it was shown that knockdown of eIF3a caused visual decreases of colonies in both cell lines. (C) By counting the colonies numbers, it was further shown that more than 60% of SW1990 colonies and 75% of Capan-1 colonies were suppressed upon eIF3a shRNA transfection. Data were obtained in triplicate with each experiments repeated three times. *P < 0.05; †P < 0.01

• 

  Fig. 4 Knockdown of eIF3a inhibited the wound recovery in SW1990 cells and Capan-1 cells. Both SW1990 cells and Capan-1 cells were subject to wound-healing assays after transfection with scramble or specific shRNA against eIF3a. Twenty-four hours after the scratch, wound recovery rates were photographed and the recovered areas which represented cell migration were quantified and averaged from three independent assays. *P < 0.01

• 

  Fig. 5 Knockdown of eIF3a decreased cell migration and invasion abilities in human pancreatic cancer cell lines. (A) SW1990 cells and Capan-1 cells were subject to transwell assays after cells were depleted of eIF3a by shRNA. Visually, the transmigrated cells were significantly decreased in the shRNA group relative to control group. (B) After counting the transmigrated cells, it was shown that the migration rate was inhibited by 55% for SW1990 cells and 46% for Capan-1 cells. Meanwhile, cell invasion rate was decreased by more than 60% for both cell lines. Each assay was repeated for three times. *P < 0.01

• 

  Fig. 6 Knockdown of eIF3a inhibited the tumorigenic ability in a xenotransplanted model. (A) Capan-1 cells with or without eIF3a depletion were subcutaneously injected into the nude mice (n = 5 for each group) to determine the impact of eIF3a depletion on pancreatic tumor growth in vivo. Tumor volumes from two groups of mice were monitored for a consecutive 4 weeks. (B) After four weeks, tumors were dissected. (C) Tumor weights from each group were weighed. It was shown that the average tumor weight from eIF3a-depleted group was significantly decreased as compared with that in the control group. *P < 0.05

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